Axon Initial Segment Loss is not Observed in the Hippocampus of a Experimental Autoimmune Encephalomyelitis Mouse Model

The axon initial segment (AIS) is fundamental for neuronal communication and action potential initiation, a characteristic which has been shown to be disrupted in inflammatory diseases such as Multiple Sclerosis (MS). Previous work from our lab has shown AIS breakdown in layer 5 of the cortex in a mouse model of MS known as experimental autoimmune encephalomyelitis (EAE). Moreover, it was shown that AIS breakdown was independent of demyelination but temporally correlated with microglial inflammatory reactivity. In order to determine if this pathology is specific to the cortex or affects other regions of the brain, we exploited these EAE induced mice and investigated AIS integrity in the hippocampus, a region associated with cognitive dysfunction in inflammatory diseases. Additionally, we used a second model of microglial activation that was shown to have AIS breakdown in the cortex. This model relies on the injection of lipopolysaccharide (LPS). Although LPS activates microglia, there is little to no direct effect on other immune cells, which is not the case for EAE. To test AIS stability in the hippocampus, EAE was induced in 12 week old c57bl/6 mice; LPS was injected into 11-12 week old c57bl/6 mice. AnkyrinG, which is an essential cytoskeletal scaffolding protein necessary for proper AIS structure and function, was used to examine AIS integrity through immunohistochemistry (IHC) combined with laser scanning confocal microscopy. Qualitative analysis of AIS produced by confocal imaging, displayed no prevalent signs of AIS shortening in early or late stages of inflammation in the EAE mouse model. The breakdown of βIV spectrin, a cytoskeletal protein that is also clustered in the AIS and is known to link ankyrinG-NaV to the actin cytoskeleton, was correlated with structural disruption of AIS. βIV spectrin breakdown products can be assessed by western blot analysis; however, no breakdown products were observed from the hippocampus of either LPS injected or EAE induced mice. These findings are preliminary but they indicate that the AIS has differential stabilities throughout the CNS, which may provide a tentative explanation for regional differences within the brain. Future studies will include quantification of microglial activation; we believe the depletion of microglial cells play a vital role in both maintaining AIS stability and AIS disruption. These cells are enigmatic and are known to play different roles in different regions of the brain. We are also interested in isolating these cells from the hippocampus and from the cortex in order to compare their expression profiles. Findings from these studies should shed light on the role microglia play in different brain regions during disease.https://scholarscompass.vcu.edu/uresposters/1227/thumbnail.jp

Bacteriophage DNA glucosylation impairs target DNA binding by type I and II but not by type V CRISPR-Cas effector complexes

Prokaryotes encode various host defense systems that provide protection against mobile genetic elements. Restriction-modification (R-M) and CRISPR-Cas systems mediate host defense by sequence specific targeting of invasive DNA. T-even bacteriophages employ covalent modifications of nucleobases to avoid binding and therefore cleavage of their DNA by restriction endonucleases. Here, we describe that DNA glucosylation of bacteriophage genomes affects interference of some but not all CRISPR-Cas systems. We show that glucosyl modification of 5-hydroxymethylated cytosines in the DNA of bacteriophage T4 interferes with type I-E and type II-A CRISPR-Cas systems by lowering the affinity of the Cascade and Cas9-crRNA complexes for their target DNA. On the contrary, the type V-A nuclease Cas12a (also known as Cpf1) is not impaired in binding and cleavage of glucosylated target DNA, likely due to a more open structural architecture of the protein. Our results suggest that CRISPR-Cas systems have contributed to the selective pressure on phages to develop more generic solutions to escape sequence specific host defense systems

Characterizing a thermostable Cas9 for bacterial genome editing and silencing

CRISPR-Cas9-based genome engineering tools have revolutionized fundamental research and biotechnological exploitation of both eukaryotes and prokaryotes. However, the mesophilic nature of the established Cas9 systems does not allow for applications that require enhanced stability, including engineering at elevated temperatures. Here we identify and characterize ThermoCas9 from the thermophilic bacterium Geobacillus thermodenitrificans T12. We show that in vitro ThermoCas9 is active between 20 and 70 °C, has stringent PAM-preference at lower temperatures, tolerates fewer spacer-protospacer mismatches than SpCas9 and its activity at elevated temperatures depends on the sgRNA-structure. We develop ThermoCas9-based engineering tools for gene deletion and transcriptional silencing at 55 °C in Bacillus smithii and for gene deletion at 37 °C in Pseudomonas putida. Altogether, our findings provide fundamental insights into a thermophilic CRISPR-Cas family member and establish a Cas9-based bacterial genome editing and silencing tool with a broad temperature range

No. 14: The State of Household Food Security in Bangalore, India

This report presents and analyzes the findings of a household food security survey conducted by the IIHS and the Hungry Cities Partnership in Bangalore, India, from April to September 2016. Surie and Sami (2017) provide essential contextual background for this report on Bangalore’s history, demography, economy, and changing food system. This report describes the survey and presents and discusses its findings. It then analyzes the food security situation and food system functions in Bangalore. The report thus provides solid background information for future research on Bangalore’s food system and lays the foundation for comparative studies with the other cities of the Hungry Cities Partnership project. The report first provides an overview of the sampling strategies for the city-wide household survey in Bangalore. It then profiles the surveyed households in terms of their demographic characteristics, economic data, livelihoods and occupations, poverty indicators, and use of social grants. The following section discusses the prevalence of food insecurity in Bangalore using various food insecurity measurements. It also explores the factors affecting food security, the impact of food price changes on food accessibility, and the relationship between food security and household characteristics. The report then examines Bangalore’s food system through people’s usage of various food sources, what foods they buy, and how they perceive supermarkets and urban agriculture

Molecular authentication of green algae Caulerpa (Caulerpales, Chlorophyta) based on ITS and tufA genes from Andaman Islands, India

109-114Indigenous and non-indigenous invasive algal species introduction or prevalence is one of the major concerns to protect the native coastal environment. Globally, several studies have reported the effect of invasive alga Caulerpa on coral reefs. To establish the genetic variation between indigenous and non-indigenous invasive species, attempts have been made to develop molecular identification of Caulerpa algal species available at the Andaman Islands. In this study, 7 visually and morphologically different species belonging to the genus Caulerpa (Chlorophyta) were collected from the intertidal regions of South and Little Andaman Islands, India. The specimens were preliminarily identified based on the morphological characters and genetically mapped using ITS2 and chloroplast tufA gene markers. Six species of the Caulerpa viz. Caulerpa racemosa, C. racemosa var lamourouxii, C. racemosa var macrophysa, C. serrulata, C. fergusonii and C. microphysa were identified using ITS2 gene, and. C. mexicana var pluriseriata was identified using tufA gene. Two varieties, C. mexicana var. pluriseriata and C. racemosa var lamourouxii were found to be invasive to Indian waters. These were earlier reported in Red sea and in Phillipine waters in the pacific ocean. Further studies are needed to elucidate the genetic divergence of the Caulerpa species present in Andaman waters using different molecular markers

Coupling immunity and programmed cell suicide in prokaryotes: Life-or-death choices

Host-pathogen arms race is a universal, central aspect of the evolution of life. Most organisms evolved several distinct yet interacting strategies of anti-pathogen defense including resistance to parasite invasion, innate and adaptive immunity, and programmed cell death (PCD). The PCD is the means of last resort, a suicidal response to infection that is activated when resistance and immunity fail. An infected cell faces a decision between active defense and altruistic suicide or dormancy induction, depending on whether immunity is “deemed” capable of preventing parasite reproduction and consequent infection of other cells. In bacteria and archaea, immunity genes typically colocalize with PCD modules, such as toxins-antitoxins, suggestive of immunity-PCD coupling, likely mediated by shared proteins that sense damage and “predict” the outcome of infections. In type VI CRISPR-Cas systems, the same enzyme that inactivates the target RNA might execute cell suicide, in a case of ultimate integration of immunity and PCD

Isolation and cellular fatty acid profile analyzation of two marine bioluminescent bacteria

192-195Two luminescent bacterial strains KOOS1 and KOOS2 isolated from surface mucus of Octopus sp. collected from Andaman were identified by their cellular fatty acid composition analyzation with the help of Microbial Identification system (MIDI). SIM indexes obtained for these isolated strains were 0.772 (KOOS1) and 0.754 (KOOS2) respectively and were identified as Photobacterium damselae and Vibrio fischeri. Major fatty acids found in Photobacterium damselae were Saturated: Dodecanoic acid (C12:0), Tetradecanoic acid (C14:0), Pentadecanoic acid (C15:0), Hexadecanoic acid (C16:0), Heptadecanoic acid (C17:0) and Octadecanoic acid (C18:0); and Unsaturated: 3-hydroxy-9-methyl decanoic acid (C11:0iso 3OH), 3-hydroxydodecanoic(C12:0 3OH), C16:1ω5c, Oleic acid (C18:1ω9c) and C18:1ω5c.In Vibrio fischeri Saturated: C12:0, Tridecanoic acid (C13:0), C15:0, C16:0, C17:0 and C18:0; and Unsaturated: C11:0iso 3OH,2-hydroxydodecanoic (C12:0 2OH), C12:03OH, C13:0iso, C14:0iso, C15:0iso, C15:0anteiso, C16:0iso, C17:0iso, C16:1ω5c, C15:0iso3OH, C17:1 ω8c and C17:1ω6c were found. Cyclopropane acids have not been detected in both Photobacterium damselae and Vibrio fischeri

Non-species specific composition of bioluminescent bacteria in non-bioluminescent squid Sepioteuthis lessoniana (Lesson, 1830)

975-981The present study investigated luminous bacterial species composition from non-bioluminescent big fin reef squid, Sepioteuthis lessoniana for the first time and revealed a new insight on the occurrence of five distinct luminous bacterial species such as Vibrio harveyi, V. campbellii, V. vulnificus, V. alginolyticus and Photobacterium damselae indicating the association of non-species specific luminous bacterial composition in S. lessoniana. Fourteen potential luminescent bacterial strains were isolated from S. lessoniana and identified based on morphological and biochemical tests. Majority of the isolates had produced industrially important extracellular enzymes- protease, chitinase and lipases. Antibiogram assay revealed that majority of the isolates were sensitive to various antibiotics and resistant to Ampicillin and Penicillin. While isolates CS1S and CS5S were sensitive to Ampicillin and Penicillin; and CS4G was sensitive to Penicillin. The present study is the first report to demonstrate luminous bacterial species diversity and extracellular enzyme production capabilities, and their antibiogram profile from big fin reef squid S. lessoniana

GC-MS analysis and antibacterial properties of the selected soft corals from South Andaman, India

No abstrac