Microbiology Commons^™

Botanical Inventory Highlights And Management Considerations For Covenant Park, David Warners, Garrett Crow, Haley Weesies

Faculty and Professional Research

In the spring and summer of 2021, Calvin University students and faculty conducted a botanical inventory and assessment of the natural areas in Covenant Park, located at 3724 Shaffer Avenue SE, Kentwood, MI 49512. Given that the property was recently acquired by the City of Kentwood Parks & Recreation, the purpose of this inventory is to inform Kentwood Parks and Recreation staff about the site’s floristic quality, diversity, and ecological significance. The specific objectives of this study are to 1) provide a comprehensive inventory of the species of plants occurring on the property, 2) identify particular areas that would benefit …

การผลิต ลักษณะสมบัติและการยับยั้งราที่ก่อโรคในพืชของสารลดแรงตึงผิวชีวภาพจาก Bacillus Licheniformis F2.2, สิริตา เสียมไหม

Chulalongkorn University Theses and Dissertations (Chula ETD)

งานวิจัยนี้ศึกษาความสามารถในการยับยั้งราก่อโรคในพืช การผลิตสารลดแรงตึงผิวชีวภาพ และผลของปัจจัยทางกายภาพที่มีต่อประสิทธิภาพในการยับยั้งราก่อโรคในพืชของสารลดแรงตึงผิวชีวภาพจาก Bacillus licheniformis F2.2 รวมถึงหาภาวะที่เหมาะสมสำหรับการผลิตสารลดแรงตึงผิวชีวภาพ จากนั้นทำบริสุทธิ์ และศึกษาลักษณะสมบัติเบื้องต้นของสารลดแรงตึงผิวชีวภาพ แล้วทดสอบการประยุกต์ใช้เพื่อยับยั้งการเกิดโรคใบไหม้ในต้นข้าว ผลการวิจัยพบว่า B. licheniformis F2.2 สามารถยับยั้งราก่อโรคในพืชได้ทั้ง 7 ชนิด ซึ่งได้แก่ Acremonium furcatum, Colletotrichum gloeosporioides, Fusarium moniliforme, Fusarium proliferatum, Fusarium solani, Pyricularia oryzae และ Phytophthora palmivora โดยมีเปอร์เซ็นต์การยับยั้งอยู่ระหว่าง 34.79±6.29-44.51±1.29% เมื่อทดสอบสมบัติในการผลิตสารลดแรงตึงผิวชีวภาพ โดยทดสอบแอกทิวิตีการสลายเม็ดเลือดแดง, การยุบตัวของน้ำเลี้ยงเซลล์, การกระจายตัวของน้ำมัน และการเกิดอิมัลชัน พบว่าส่วนใสของน้ำเลี้ยงเชื้อของ B. licheniformis F2.2 ให้ผลบวกกับทุกการทดสอบ ซึ่งแสดงว่า B. licheniformis F2.2 สามารถผลิตสารลดแรงตึงผิวชีวภาพได้ จากการศึกษาผลของอุณหภูมิและความเป็นกรดเบสต่อประสิทธิภาพในการยับยั้งราก่อโรคในพืชของส่วนใสของน้ำเลี้ยงเชื้อของ B. licheniformis F2.2 พบว่าเมื่อผ่านการบ่มน้ำเลี้ยงเชื้อที่ 20-121 องศาเซลเซียส และที่ pH 2-12 พบว่าสมบัติการเกิดอิมัลชันของสารลดแรงตึงผิวชีวภาพก็ยังมีความเสถียร และประสิทธิภาพการยับยั้งราก่อโรคในพืชทั้ง 7 ชนิดยังคงอยู่ โดยสามารถยับยั้ง P. oryzae ซึ่งเป็นสาเหตุของโรคไหม้ในข้าวได้มากที่สุด จากการแปรผันแหล่งคาร์บอนและแหล่งไนโตรเจนพบว่าอาหาร MOLP ที่มีซูโครสเป็นแหล่งคาร์บอน, แอมโมเนียมคลอไรด์และสารสกัดจากยีสต์เป็นแหล่งไนโตรเจนส่งเสริมการเจริญเติบโตของ B. licheniformis F2.2 ได้ดีที่สุด และเหมาะสมต่อการผลิตสารลดแรงตึงผิวชีวภาพที่มีฤทธิ์ยับยั้งราก่อโรคในพืช โดยมีค่าดัชนีการเกิดอิมัลชันที่ 24 ชั่วโมง (E24) 62.92±2.30% สำหรับความสัมพันธ์ระหว่างระยะเวลาการเจริญกับการผลิตสารลดแรงตึงผิวชีวภาพที่มีฤทธิ์ยับยั้งราก่อโรคในพืช พบว่าเมื่อเลี้ยง B. licheniformis F2.2 ในอาหาร MOLP พบว่ามีความสามารถในการยับยั้งราก่อโรคในพืชทั้ง 7 ชนิดสูงสุดเมื่อแบคทีเรียเข้าสู่ stationary phase ที่ 24 ชั่วโมง เมื่อสกัดสารลดแรงตึงผิวชีวภาพด้วยเมธานอล 100% (ปริมาตร/ปริมาตร) …

Detection And Potential Virulence Of Viable But Non-Culturable (Vbnc) Listeria Monocytogenes: A Review, Nathan E. Wideman, James D. Oliver, Philip Glen Crandall, Nathan A. Jarvis

Food Science Faculty Publications and Presentations

The detection, enumeration, and virulence potential of viable but non-culturable (VBNC) pathogens continues to be a topic of discussion. While there is a lack of definitive evidence that VBNC Listeria monocytogenes (Lm) pose a public health risk, recent studies suggest that Lm in its VBNC state remains virulent. VBNC bacteria cannot be enumerated by traditional plating methods, so the results from routine Lm testing may not demonstrate a sample’s true hazard to public health. We suggest that supplementing routine Lm testing methods with methods designed to enumerate VBNC cells may more accurately represent the true level of risk. This review …

The Gut Microbiome Of Hapalemur Griseus Griseus, Propithecus Diadema, And Propithecus Coquereli: A Comparative Study Between Wild And Captive Lemur Species, Brenna Ritchie

Graduate Research Theses & Dissertations

Madagascar is home to thousands of endemic species, making it a biodiversity hotspot that is rapidly losing its fauna due to deforestation and human expansion. Efforts to restore and preserve this unique island are ongoing and several researchers have placed their focus on preventing species extinctions. One research avenue that can help contribute to these efforts is to study health and wellness of animals located in areas affected by these changes. In this study, I sought to characterize microbial community composition of two native lemur species found in Madagascar and compare those profiles to similar species found in captivity in …

Narratives On The Academic Resilience Of Former Homeless Students, Alia Aboulhosn

Walden Dissertations and Doctoral Studies

The number of homeless students in U.S public schools continues to rise. Although many homeless students drop out due to challenges they face, lacking is research on how some homeless students overcome challenges and graduate high school, referred to as academically resilient. The purpose of this qualitative narrative study was to gain a deeper understanding of the experiences that promoted former students who were homeless during their senior year of high school to graduate despite adversity, using Resiliency Theory as the theoretical framework. Semi-structured interviews with five participants provided insight into the broader sociopolitical and personal contexts in their stories, …

Investigating The Roles Of Lipids In Staphylococcus Aureus Infection, Xi Chen

Dissertations

The Gram-positive bacterium Staphylococcus aureus is a major threat to public health and causes a multitude of infections, ranging from mild skin and soft infection, to more severe diseases including sepsis, osteomyelitis and infective endocarditis. As a successful pathogen, S. aureus employs various mechanisms to invade host tissues, evade immune responses, and survive in the host environment. One critical adaptive trait of S. aureus that promotes virulence and survival is the ability to maintain membrane homeostasis via flexible modifications to its lipid composition in response to the surrounding lipid environment. Phospholipids are the major component of the bacterial membrane and …

F Plasmids In Escherichia Coli Decrease Permissivity To Coliphage, Cesar Eber Montelongo Hernandez

Dissertations

The urinary tract contains a community of bacteria called the urinary microbiota (urobiota) thatmay be relevant to health; the genomic component of the urobiota is the urinary microbiome (urobiome). Urinary bacteria have been associated with both asymptomatic states and disease conditions, such as urinary tract infection (UTI), overactive bladder (OAB), and urge urinary incontinence (UUI). Some bacteria, such as E. coli, are considered urinary pathogens (uropathogens) but also can be commensals. Bacteriophage (phage) are ubiquitous in nature and likely shape bacterial populations in every niche; thus, phage may be one factor that modulates the urobiota. Phages have a specific host …

Identification And Characterization Of Calcium As A Biofilm Promoting Signal By Vibrio Fischeri, Alice Hannah Tischler

Dissertations

The marine bacterium, V. fischeri actively engages in an exclusive partnership in the light organ of its squid host, Euprymna scolopes. A critical step in this colonization process is the formation of a bacterial aggregate, or biofilm, which is a community of bacteria embedded within a protective extracellular matrix. While V. fischeri readily forms biofilms in nature, genetic overexpression must be utilized to achieve biofilm formation in the laboratory. Recent work investigating media composition led myself and others to evaluate how seawater salts impacted growth and biofilm formation in a number of mutant backgrounds, leading to the identification of calcium …

Assessing The Role Of The Interaction Between Polyamines And Heparan Sulfate In Coxsackievirus B3 Viral Infection, Oreoluwa Omoba

Master's Theses

Non-polioenteroviruses are highly infectious viruses that typically cause mild asymptomatic cases but can cause severe disease such as aseptic meningitis and dilated cardiomyopathy. Coxsackievirus B3 (CVB3) is a member of this group and is prevalent in the United States and Eastern Asia. Viruses like CVB3 rely on the host cell for many molecules in order to replicate. One such set of molecules is the polyamines. Polyamines are small, positively charged molecules that play a role in a multitude of cellular processes including RNA/DNA stabilization, gene expression, translation, and regulating membrane fluidity. Previous studies have shown CVB3 relies on polyamines for …

Elucidating Regulatory Connections Between The Second Messenger Cyclic-Di-Gmp With Biofilm And Motility Phenotypes Of Vibrio Fischeri, Ali Razvi

Master's Theses

The symbiosis between the gram-negative marine bacterium Vibrio fischeri and its host Euprymna Scolopes occurs when the bacteria reaches and colonizes the light organ. This pathway to colonization begins in the outer marine environment and finishes within the deep crypts of the light organ, with states of biofilm formation and dispersal in between. This switch between biofilm formation to dispersal is important as the bacteria transitions from the surface on the light organ into a pore leading to the interior of the light organ. This dispersal is regulated by LAP system which is dependent on the second messenger c-di-GMP. Through …

Comparison Of The Effects Of Land Use / Land Cover On Water Quality And The Intertidal Marine Macroalgae Communities At The Fluvial Outflows Of The Dingle Peninsula, Cassandra Lew

Theses

The Milltown, Feohanagh, and Owenmore river basins located on the Dingle Peninsula in Ireland were studied to see how variation in land use and land cover (LU/LC) affected water quality and the macroalgal communities at each river’s respective fluvial outflow. Abundance and composition of macroalgae in river and coastal water bodies have been designated as an indicator of water quality according to the Water Framework Directive 2000. The results of this study found that the Feohanagh’s macroalgal community had the highest species richness for most months while the Milltown had the highest Shannon Diversity Index. The Owenmore’s community was consistently …

Free-Living Diazotrophs And The Nitrogen Cycle In Natural Grassland Revealed By Culture Dependent And Independent Approaches, Amrit Koirala

Electronic Theses and Dissertations

Biological nitrogen fixation contributes to half of the global supply of nitrogen to the biosphere. It is carried out by a diverse group of prokaryotes called diazotrophs via the nitrogenase enzyme. Nitrogen fixation research is focused on the narrow group of symbiotic diazotrophs, and the vast majority of free-living diazotrophs which contribute significantly to fixed nitrogen are yet to be explored. The goal of this research was to access phylogeny of diazotrophs considering the most up-to-date genomic information and apply that knowledge to understand the diversity of free-living diazotrophs in a natural grassland ecosystem, both by culture dependent and independent …

A Survey Of Opportunistic Fungi In The Portsmouth, Nh Region, Chloe E. Richard

Honors Theses and Capstones

Opportunistic mycoses are caused by ubiquitous fungi that we encounter daily. This study identified the most common opportunistic fungi in a local area during the current winter season. A survey of fungi was conducted in the Portsmouth, New Hampshire region by taking environmental samples from both indoor and outdoor locations. The samples were then cultured and identified using colony morphology, microscopic characteristics, and biochemical testing. Despite the cold weather, the results showed that fungal species are still prevalent outdoors, with the most variation in single samples coming from areas with exposed soil and decaying vegetation, while less fungal presence was …

Roles Of Non-Frankia Bacteria In Root Nodule Formation And Function In Alnus Sp., Kelsey Christine Mercurio

Honors Theses and Capstones

Plant roots are home to a wide variety of beneficial microbes; understanding and optimizing plant-microbe interactions may be critical to enhance global food security in a sustainable, equitable way. With the help of their nitrogen-fixing bacterial partner, Frankia, actinorhizal plants form symbiotic root nodules and play important roles in agroforestry and land reclamation. However, Frankia does not live alone in nodules, and the other microbial residents may contribute to nodule formation and function. We collected root nodules from alder trees (Alnus sp.) in 2018 and 2019, then isolated DNA and individual bacterial strains to characterize the nodule microbial …

Predicting, Monitoring And Controlling The Growth Of Listeria In Complex Food Matrices, Laura Mary Nyhan

Theses

Listeria monocytogenes is a foodborne pathogen which is a significant challenge in food production, particularly for ready-to-eat (RTE) products. Incidence of Listeria in RTE foods can be reduced by the application of intelligent combinations of preservative factors or hurdles, while data quantifying the efficacy of hurdle combinations can be exploited and used in the area of predictive microbiology. Traditional culture-based techniques, such as viable plate counts, are commonly used to monitor the growth/survival of microorganisms in foods, however these methods are tedious, time-consuming and due to their destructive nature, are associated with low reproducibility and high variability. Therefore, rapid, non-invasive …

Global Coordination Between Gene Expression Processes In C. Crescentus, James Raymond Aretakis

Wayne State University Dissertations

Gene expression is an essential process of all organisms which occurs in two steps, mRNA transcription, and translation and which is shut off by mRNA decay. For a bacterium to grow and divide, these steps need to be tuned to optimize the synthesis rates of thousands of proteins from their genes. Genome-wide experiments were used to determine the absolute transcription rates, mRNA degradation rates, and translation rates on a gene by gene basis in the model bacterium Caulobacter crescentus. This not only provides a system biology view of the organism’s gene expression resource allocation, but also provides a chance to …

A Review Of The Molecular Mediators Of Host-Baculovirus Interactions In Lepidoptera, Olivia Crouse

Honors Program Theses

Iowa farmlands occupy approximately 85% of its 35.7 million acres, and Iowa leads the United States in corn and soybean production, with an estimated 2.58 billion and 502 million bushels of corn and soybean produced in 2019, respectively. This successful trend in annual yields has span forty plus years and been consequential to the simultaneous implementation of agricultural- and science-based practices. Genetically modified corn and soybean crop varieties express genes from Bacillus thuringiensis (Bt) which encode insecticidal proteins that protect, with high specificity, against insect pests. While researchers continue to study the molecular mechanism(s) of Bt extensively, alternative environmentally safe …

Bacillus Bacteriophage Q11 May Have A 5' Covalently Attached Protein, Emily Cornelius

Honors Program Theses

Bacteriophages (phages) are viruses that specifically parasitize on bacteria and are found almost everywhere. Phages Phi 29 and Q11 are both small phages found in soil that attack Bacillus hosts. Previous Phi 29 studies verified a covalently bonded protein at the 5’ end of its double-stranded linear DNA. Q11 may have a similar 5’ terminal protein, which blocks complete sequencing of that DNA region. Because Q11 and phi29 are both small short-tailed phages that infect similar hosts, it could be possible the proteins are structurally similar. In previous work, I have attempted to use restriction endonuclease and exonuclease enzyme digests …

Evaluation Of Archeal Growth On Complex Media And Antibiotic Regimes, Melanie Miller

Honors Program Theses

The first Archaea that were studied by scientists came from extreme environments such as fissures at the bottom of oceans where hydrothermal vents formed. It was perhaps surprising that later sampling of environmental DNA revealed Archaea inhabit normal environments as well, for example they are globally distributed in soils. These mesophilic soil species were only recently grown in laboratory pure cultures. In this study, we will expand on this work by feeding the community of soil microbes with various dilutions of tryptic soy broth and other complex media in combination with an antibiotic regime. This will allow us to compare …

The Search For Life In Extreme Environments: Measuring Diversity And Quantity Of Archaea In Wind Cave National Park, Nicole Geerdes

Honors Program Theses

Archaea have the potential to play an essential role in their environment at the base of the food chain. In particular, most of the known species in the phylum Thaumarchaea are autotrophs that can fix their own carbon using energy from ammonia oxidation, that is, they produce their own organic molecules from carbon dioxide by burning nitrogen-based fertilizer. Since their discovery, it has been difficult to learn more about these microbes because they do not grow well in pure laboratory culture. Therefore, much of what is known about them comes from analyzing environmental DNA samples. This allowed researchers to discover …

High Prevalence Of Pre-Existing Serological Cross-Reactivity Against Severe Acute Respiratory Syndrome Coronavirus-2 (Sars- Cov-2) In Sub-Saharan Africa, For Yue Tso, Salum Lidenge, Phoebe B. Peña, Ashley A. Clegg, John R. Ngowi, Julius Mwaiselage, Owen Ngalamika, Peter Julius, John T. West, Charles Wood

Nebraska Center for Virology: Faculty Publications

Objective: Significant morbidity and mortality have occurred in the USA, Europe, and Asia due to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), whereas the numbers of infections and deaths in sub-Saharan Africa (SSA) have remained comparatively low. It has been hypothesized that exposure of the population in SSA to other coronaviruses prior to the COVID-19 pandemic resulted in some degree of cross-protection against SARS-CoV-2 infection and pathogenesis. We evaluated this hypothesis by comparing SARS-CoV-2 cross-reactive antibodies in pre-pandemic plasma samples collected from SSA and the USA.

Method: Pre-COVID-19 pandemic plasma samples from SSA and the USA were collected and tested by …

The 20th Anniversary Meeting Of The Rocky Mountain Virology Association, Joel Rovnak, Laura A. St Clair, Carley Mcalister, Chinemerem P. Ogbu, Jessica Smolenske, Randall J. Cohrs, Rushika Perera

Nebraska Center for Virology: Faculty Publications

Due to the COVID-19 pandemic and multiple devastating forest fires, the 2020 meeting of the Rocky Mountain Virology Association was held virtually. The three-day gathering featured talks describing recent advances in virology and prion research. The keynote presentation described the measles virus paradox of immune suppression and life-long immunity. Special invited speakers presented information concerning visualizing antiviral effector cell biology in mucosal tissues, uncovering the T-cell tropism of Epstein-Barr virus type 2, a history and current survey of coronavirus spike proteins, a summary of Zika virus vaccination and immunity, the innate immune response to flavivirus infections, a discussion concerning prion …

Chlorovirus Pbcv-1 Multidomain Protein A111/114r Has Three Glycosyltransferase Functions Involved In The Synthesis Of Atypical N-Glycans, Eric Noel, Anna Notaro, Immacolata Speciale, Garry A. Duncan, Cristina De Castro, James L. Van Etten

Nebraska Center for Virology: Faculty Publications

The structures of the four N-linked glycans from the prototype chlorovirus PBCV-1 major capsid protein do not resemble any other glycans in the three domains of life. All known chloroviruses and antigenic variants (or mutants) share a unique conserved central glycan core consisting of five sugars, except for antigenic mutant virus P1L6, which has four of the five sugars. A combination of ge- netic and structural analyses indicates that the protein coded by PBCV-1 gene a111/114r, conserved in all chloroviruses, is a glycosyltransferase with three putative domains of approximately 300 amino acids each. Here, in addition to in silico sequence …

Strategies Targeting Hemagglutinin As A Universal Influenza Vaccine, Brianna L. Bullard, Eric A. Weaver

Nebraska Center for Virology: Faculty Publications

Influenza virus has significant viral diversity, both through antigenic drift and shift, which makes development of a vaccine challenging. Current influenza vaccines are updated yearly to in- clude strains predicted to circulate in the upcoming influenza season, however this can lead to a mismatch which reduces vaccine efficacy. Several strategies targeting the most abundant and im- munogenic surface protein of influenza, the hemagglutinin (HA) protein, have been explored. These strategies include stalk-directed, consensus-based, and computationally derived HA immunogens. In this review, we explore vaccine strategies which utilize novel antigen design of the HA protein to improve cross-reactive immunity for development …

Zika Virus Infection Induces Endoplasmic Reticulum Stress And Apoptosis In Placental Trophoblasts, Philma Glora Muthuraj, Prakask K. Sahoo, Madison Kraus, Taylor Bruett, Arun S. Annamalai, Aryamav Pattnaik, Asit K. Pattnaik, Siddappa N. Byrareddy, Sathish Kumar Natarajan

Nebraska Center for Virology: Faculty Publications

Zika virus (ZIKV) infection to a pregnant woman can be vertically transmitted to the fetus via the placenta leading to Congenital Zika syndrome. This is characterized by microcephaly, retinal defects, and intrauterine growth retardation. ZIKV induces placental trophoblast apoptosis leading to severe abnormalities in the growth and development of the fetus. However, the molecular mechanism behind ZIKV-induced apoptosis in placental trophoblasts remains unclear. We hypothesize that ZIKV infection induces endoplasmic reticulum (ER) stress in the trophoblasts, and sustained ER stress results in apoptosis. HTR-8 (HTR-8/SVneo), a human normal immortalized trophoblast cell and human choriocarcinoma-derived cell lines (JEG-3 and JAR) were …

Advancing Cyanobacteria Biomass Estimation From Hyperspectral Observations: Demonstrations With Hico And Prisma Imagery, Ryan E. O'Shea, Nima Pahlevan, Brandon Smith, Mariano Bresciani, Todd Egerton, Claudia Giardino, Lin Li, Tim Moore, Antonio Ruiz-Verdu, Steve Ruberg, Stefan G.H. Simis, Richard Stumpf, Diana Vaičiūtė

Biological Sciences Faculty Publications

Retrieval of the phycocyanin concentration (PC), a characteristic pigment of, and proxy for, cyanobacteria biomass, from hyperspectral satellite remote sensing measurements is challenging due to uncertainties in the remote sensing reflectance (∆R_rs) resulting from atmospheric correction and instrument radiometric noise. Although several individual algorithms have been proven to capture local variations in cyanobacteria biomass in specific regions, their performance has not been assessed on hyperspectral images from satellite sensors. Our work leverages a machine-learning model, Mixture Density Networks (MDNs), trained on a large (N = 939) dataset of collocated in situ chlorophyll-a concentrations (Chla), …

Biology And Molecular Biology Of Ixodes Scapularis, Daniel E. Sonenshine, Ladislav Šimo, Justin D. Radolf (Ed.), D. Scott Samuels (Ed.)

Biological Sciences Faculty Publications

This chapter describes the biology of the tick Ixodes scapularis in relation to its role as the vector of the Lyme disease agent, Borrelia burgdorferi. Following a review of the internal anatomy of the tick, we review basic molecular processes that contribute to an understanding of the dynamics of the tick's specialized parasitic processes, including attachment behavior, salivation; silencing of host anti-inflammatory responses to enable blood ingestion at the dermal feeding site; hemoglobin digestion and reproduction. The chapter is divided into three parts: 1) systematic and anatomical characteristics of ticks; 2) host finding, attachment, salivary disruption of host defenses, blood …

Modeling Stromatolite Formation With Diffusion-Limited Aggregation, Laura E. Stevens

Graduate Student Theses, Dissertations, & Professional Papers

Stromatolites, microbialites, and other microbially induced sedimentary structures exist in the rock record as far back as 3.6 billion years ago and continue to form in the present day. Better characterizing these structures and better understanding how they form is crucial in distinguishing these biosignatures from similar, abiotic structures, which can help us to understand the conditions of early Earth and early Mars. To that end, I have modified DLA 3D EXT, an open-source stromatolite modeling program, to more closely reflect the process of microbial trapping-and-binding by filamentous microbes in a calcite-precipitating hot spring system. This modified program includes a …

การยับยั้งการติดไวรัสหัวเหลืองในกุ้งด้วยอนุภาคเสมือน Penaeus Stylirostris เดนโซไวรัสกับอาร์เอ็นเอสายคู่ของไวรัส, จารุวรรณ วรวิทยธาดา

Chulalongkorn University Theses and Dissertations (Chula ETD)

ไวรัสหัวเหลืองเป็นไวรัสก่อโรคที่ทำให้กุ้งหลายชนิดตายเป็นจำนวนมากในระยะเวลาสั้น ส่งผลให้อุตสาหกรรมการเพาะเลี้ยงและส่งออกกุ้งของไทยเกิดความเสียหายเป็นอย่างมาก ในปัจจุบันมีการใช้เทคโนโลยีทางอณูชีววิทยา นำอาร์เอ็นเอสายคู่ (dsRNA) มากระตุ้นให้เกิดกระบวนการ RNA interference (RNAi) เพื่อควบคุมและยับยั้งการติดไวรัสในกุ้ง โดยยีนเป้าหมายที่มีประสิทธิภาพในการยับยั้งไวรัสหัวเหลือง คือ ยีน protease (pro) อย่างไรก็ตามการขนส่งอาร์เอ็นเอสายคู่ก็ยังคงมีข้อจำกัด เปลือกหุ้มไวรัสบางชนิดสามารถรวมตัวเกิดเป็นอนุภาค-เสมือนไวรัส (Virus-like particle) สามารถใช้เป็นตัวขนส่งที่มีประสิทธิภาพและเข้าจับกับเซลล์เจ้าบ้านอย่างจำเพาะเจาะจงได้ โดยก่อนหน้านี้ได้มีการแสดงออกร่วมกันของยีนโปรตีนเปลือกหุ้มไวรัสของ Penaeus stylirostris เดนโซไวรัส (PstDNV) และอาร์เอ็นเอสายคู่ที่จำเพาะต่อยีนของไวรัสภายในเซลล์แบคทีเรียเดียวกัน พบว่าสามารถยับยั้งการเพิ่มจำนวนของไวรัสหัวเหลืองในกุ้งได้ แต่ทว่ากระบวนการแสดงออกร่วมกันนี้ ไม่สามารถควบคุมระดับการแสดงออกของยีนทั้งสองได้ ดังนั้นวัตถุประสงค์ของงานวิจัยนี้ คือ สร้างอนุภาคเสมือนไวรัส PstDNV กับอาร์เอ็นเอสายคู่ที่จำเพาะต่อยีน pro ของไวรัสหัวเหลือง (PstDNV VLPs-dspro) ให้อยู่ภายใต้โปรโมเตอร์เดียวกัน เพื่อนำไปประยุกต์ใช้และเพิ่มประสิทธิภาพในการป้องกันการติดเชื้อไวรัสหัวเหลืองในกุ้ง โดยการโคลนอาร์เอ็นเอสายคู่ที่จำเพาะต่อยีน pro ของไวรัสหัวเหลืองเข้าสู่เวกเตอร์สำหรับการแสดงออกที่มียีนโปรตีนเปลือกหุ้มไวรัส PstDNV เพื่อควบคุมการแสดงออกของยีนทั้งสองภายใต้โปรโมเตอร์ต่อ T7 RNA polymerase จากนั้นแสดงออกใน E. coli Rosetta-gami โดยผ่านการชักนำด้วย IPTG ที่ความเข้มข้นสุดท้าย 0.4 มิลลิโมลาร์ ที่อุณหภูมิ 30 องศาเซลเซียส เป็นเวลา 3 ชั่วโมง จากผลการวิเคราะห์พบทั้งรีคอมบิแนนท์โปรตีนขนาดประมาณ 37 กิโลดาลตัน ที่เข้าจับกับแอนติบอดีที่จำเพาะต่อ Histidine ได้ และพบอาร์เอ็นเอสายคู่ที่จำเพาะต่อยีน pro ของไวรัสหัวเหลือง ขนาดประมาณ 400 คู่เบส จากนั้นรีคอมบิแนนท์โปรตีนถูกทำให้บริสุทธิ์ด้วย Affinity chromatography และตรวจสอบภายใต้กล้องจุลทรรศน์อิเล็กตรอนแบบส่องผ่าน (TEM) พบอนุภาค-เสมือนไวรัส PstDNV ที่มีการกระจายตัวเป็นอนุภาคเสมือนไวรัสเดี่ยว คล้ายกับอนุภาคเสมือนไวรัส PstDNV ที่พบในธรรมชาติ ซึ่งต่างจากอนุภาคเสมือนไวรัส PstDNV ที่ได้จากการแสดงออกร่วมกันของยีนโปรตีนเปลือกหุ้มไวรัส PstDNV กับอาร์เอ็นเอสายคู่ที่จำเพาะต่อยีนของไวรัสหัวเหลืองภายในเซลล์แบคทีเรียเดียวกัน (PstDNV VLPs-dspro co-expression) เกิดการจับกันของอนุภาคเสมือนไวรัส เมื่อตรวจสอบอาร์เอ็น-เอสายคู่จากอนุภาคเสมือนไวรัส PstDNV พบว่าอาร์เอ็นเอสายคู่นั้นถูกห่อหุ้มอยู่ทั้งภายในและอยู่ล้อมรอบอนุภาคเสมือนไวรัส PstDNV โดยพบว่ามีปริมาณอาร์เอ็นสายคู่ต่อไมโครกรัมของรีคอมบิแนนท์โปรตีนของ …

Expression Of Programmed Cell Death 1 (Pd-1) In Human Monocytic Cell Line Thp-1 And Its Regulatory Mechanism, Kittitach Sri-Ngern-Ngam

Chulalongkorn University Theses and Dissertations (Chula ETD)

Cancer is the leading cause of death worldwide. Immune system defends the host against foreign antigens, not only pathogens or their products, but also cancer cells via coordinated complex responses. The immune response against cancer; however, often poorly control the cancer growth because tumors can effectively escape the immune defense mechanisms by various immune evasion strategies. Programmed cell death 1 (PD-1) is a co-inhibitory immune checkpoint receptor expressed on various immune cells, especially in activated T cells. Engagement of PD-1 with its ligand, PD-L1 or PD-L2, leads to the negative regulation of T cell functions, resulting in impaired antitumor immunity. …