Life Sciences Commons^™

The Occurrence Of Listeria Monocytogenes In The Mushroom Production Chain And The Use Of Bacteriophage For Its Control, Vincenzo Pennone

Theses

In Agaricus bisporus production, the occurrence of L. monocytogenes is still poorly understood. A number of studies have shown the presence of L. monocytogenes in the mushroom processing environment, suggesting its source from raw materials storage areas (Viswanath et al, 2013; Murugesan et al, 2015). In this study, 10 mushroom producers and three raw material (compost and casing) producers participated in two surveys on the occurrence of L. monocytogenes in the most high-risk areas of the production environment. In the first survey, floors, drains, platforms, crates, raw materials and mushrooms were tested for L. monocytogenes presence and the results were …

Crystallographic Structure Determination Of Bacteriophage-Encoded Enzymes That Specifically Target Pathogenic Bacteria, Marta Sanz Gaitero

Theses

Antibiotic resistance is becoming a serious public health concern. Infections that some decades ago could be treated with antibiotics now sometimes do not respond to traditional treatment, causing higher mortality and economic losses. An alternative to the use of antibiotics are bacteria's natural predators, bacteriophages (or phages), and specifically their lytic enzymes. These proteins are produced by phages to degrade bacterial peptidoglycan to inject their genetic material into the bacteria (virion-associated peptidoglycan hydrolases) or to release their progeny once the infection is finished (endolysins). They can be applied exogenously to lyse Gram-positive bacteria or be genetically engineered to lyse Gram-negative …

Exploiting Bacteriophages And Associated Peptidoglycan Hydrolases With Potential For Biocontrol In Food Related Applications, Lorraine Endersen

Theses

The work presented in this thesis describes the isolation and characterisation of novel bacteriophages, and the subsequent exploitation of their bactericidal properties against two opportunistic food related pathogens, Mycobacterium avium subsp. paratuberculosis (MAP) and Cronobacter sakazakii.

MAP is a proven animal pathogen known to cause Johne’s disease in cattle but has been implicated as a causative agent of Crohn’s disease in humans. Despite the fact that this association has yet to be proven, significant focus has been directed towards evaluating the consequences of consuming milk contaminated with MAP. Accordingly, six mycobacteriophages were isolated and characterised in terms of temperature and …

Gastroenteritis Viruses In Ireland: Epidemiology And Evolution, Lynda Gunn

Theses

Viral gastroenteritis is a common disease of both humans and animals. This thesis focuses on the use of classical virology, coupled with phylogenetics to study gastroenteritis viruses [Bocavirus (BoV), Coronavirus (CoV), and Rotavirus A (RVA)], isolated from selected samples from humans and animals. RVA was associated with outbreaks in elderly care facilities in Northern Ireland, isolates were genotyped and we reported the first detection of G2 lineage lie RVA in Ireland, with other common human RVA genotypes G1P[8], G2P[4] and G9P[8]. Evolutionary dynamics of human RVA VP7 genotypes was examined using codon models of evolution, results show heterogeneous selective pressure …

The Isolation And Characterisation Of Acientobacter Baumannii Bacteriophage, Mohamed Farag M. Elbreki

Theses

Acinetobacter baumannii is a pathogen, which has an increasing prevalence of multidrug resistance. It exists widely in natural environments, and frequently in health-care settings where it has proven difficult to eradicate using antibiotic therapy. A possible alternative to conventional antibiotics is the use of bacteriophages (phages) as antibacterials. In this project, extensive screening of environmental samples such as soils, sewage and waters was undertaken to identify suitable phages. Three phages were successfully isolated from municipal waste in the Cork area. These were subsequently characterized in detail and examined by electron microscopy, which showed that they were all of the Myoviridae …

Exploitation Of The Bacteriophage-Derived Peptidase Chapk, Pierre-Mehdi Hadbi

Theses

This project investigated an anti-staphylococcal bacteriophage-derived peptidoglycan hydrolase enzyme, namely a cysteine-histidine amydohydrolase/peptidase (CHAP_k). The study focused firstly on optimizing the production of recombinant CHAP_k, which was previously cloned in an E. coli expression system, and sets out the resulting optimal Standard Operating Procedures (SOPs) for production, purification and determination of concentration and activity of the resulting protein stock. A typical yield resulting from the investigation was 300pg of CHAP_k from one litre of E.coli culture. The activity of the purified enzyme was typically in the region of 190U nmol ’. Secondly, the project investigated …

Phylogenetic Analysis Of Rotavirus Strains Gl, G3 5g9 5p[8] And P[4] Using Both Neighbour-Joining And Maximum Parsimony, Jenny Cotter

Theses

Globally rotavirus is one of the most important causes of gastroenteritis in children and affects nearly all children by the age of 5 years. It also infects adults with most severe symptoms in the elderly or immune compromised. The most common strains associated with human rotavirus gastroenteritis are G1-G4 and G9 along with P[8],[6]and [4].All previous studies used only neighbour-joining method in the phylogenetic analysis of rotavirus genes. In this study rotavirus genes VP7 and VP4 for rotavirus strains G1 .G3 ,G9 , P[8] and P[4] were analysed in MEGA4.0 using both neighbour-joining and maximum parsimony. The main difference in …

Purification And Applications Of A Bacteriophage-Derived Enzyme Chapk Against Problematic Staphylococci., Niamh Marie O'Shea

Theses

The bacteriophage K-derived peptidase CHAPk (Cysteine/Histidine-dependant amidohydrolase peptidase) can be produced from E. coli XL 1-Blue. Standard operating procedures for the production, purification and quality assessment of the peptidase were developed and set out in detail in this thesis. CHAPk exhibits strong antistaphylococcal properties and the purified enzyme was evaluated for its potential to eliminate S. aureus from the bovine udder and from human skin. As a teat dip CHAPk reduced the levels of S. aureus by three to five log cycles on both animal and human surfaces. CHAPk activity in milk was evaluated using challenge assays against the bovine …

Purification, Characterisation And Applications Of A Cloned Staphylococcal Bacteriophage Lysin For The Control Of Multi-Drug Resistant Staphylococcus Aureus Including Mrsa, Mark Fenton

Theses

The work presented in this thesis focuses on the purification, characterisation and practical applications of an anti-staphylococcal lysin (LysK) and its single-domain truncated derivative (CHAP_k) against pathogenic staphylococci including meticillin resistant Staphylococcus aureus (MRSA). Both proteins were purified to greater than 90% homogeneity. Affinity chromatography was applied as the optimum method for LysK purification and ion-exchange chromatography was successfully applied to CHAP_k. The lytic spectrum of LysK was investigated by zymogram analysis where the enzyme exhibited strong activity against a range of staphylococci regardless of their origin, sequence type (ST) or antibiotic resistant profile. This analysis …

Detection And Molecular Characterisation Of Rotavirus, Adenovirus And Bocavirus Circulating In Southern Ireland, Olivia Cashman

Theses

This study reports the detection and molecular characterisation of group A rotavirus circulating in the south of Ireland in both the bovine and human population. Characterisation of the group A rotavirus was carried out through the extraction of viral nucleic acid in faecal specimens collected from hospitals in the Cork and Waterford area and Cork regional veterinary laboratory from 2006-2009. Other gastroenteritis viruses characterised during this study include adenovirus type 41, and the newly identified gastrointestinal virus, human bocavirus, which was identified for the first time in Ireland during this study. The human bocaviruses identified in children in the south …

Detection And Molecular Characterization Of Canine Enteric Viruses In Southern Ireland., Susan Mcelligott

Theses

Enteric viruses, such as canine parvovirus (CPV) and canine coronavirus (CCoV), are a major cause of gastroenteritis in dogs. More serious clinical symptoms occur in young pups, or when both of these viruses infect an animal simultaneously. Caliciviruses and rotaviruses can also cause gastroenteritis in dogs, but these viruses, while causing significant problems in other animal species and humans, are considered to be less significant pathogens of canines. However, on occasion, these viruses have been reported to produce severe clinical symptoms such as vomiting, haemorrhagic enteritis and severe dehydration.

Many viral vaccines for canines are formulated using old vaccine viral …

Identification And Molecular Characterisation Of Mycobacteriophages And Their Lysis Proteins, Marine Henry

Theses

Given their potential as specific and natural biocontrol agents, bacteriophages and their associated proteins have become the focus of renewed interest over the last decade. This study reviews the current state of mycobacterial diseases control worldwide and presents how phage therapy and the production of recombinant bacteriophage lysis proteins can be applied as novel biocontrol strategies for mycobacteria.

Chapter 11 presents an overview of the predicted LysA and LysB proteins in all the mycobacteriophages sequenced to date and proposes an in silico 3D predictive model of the lysin B of a recently isolated and characterised mycobacteriophage. Our data also confirms …

Molecular Characterization Of Gastroenteritis Viruses Among Equines And Porcines In Southern Ireland From 2005-2007., P. J, Collins

Theses

Rotaviruses are a major cause of gastroenteritis in humans and animals. To date, seven serogroups (A to G) of rotavirus have been determined. Group A Rotaviruses (GARVs) are enteric pathogens, causing acute watery dehydrating diarrhea in various host species, including birds and mammals. Rotaviruses account for ~611,000 child deaths each year, mainly in developing countries. Likewise, rotavirus-associated enteritis is a major problem in young calves, weaning and post-weaning piglets and foals. Vaccines against the most important serologic group of rotaviruses (GARVs) are available for the prevention of rotavirus diseases in cows and horses, and, more recently, they have been made …

Detection And Molecular Epidemiological Analysis Of Acute Gastroenteritis Viruses In The South Of Ireland, Grainne Lennon

Theses

Rotavirus infections are considered to be the most common cause of severe gastroenteritis in children under the age of 5 years, and are responsible for up to 611,000 deaths annually, mainly in developing countries. From 1997 to 1999, 16 hospital laboratories in the Republic of Ireland reported to detection of 4,643 cases of rotaviral disease, while in 2004 and 2005 and increase of 1,600 and 2,251 rotavirus cases were reported. These rotavirus cases have a significant impact on the healthcare system with the minimum cost per case being €728.40. A large percentage of the infections were reported in neonates, suggesting …

Analysis Of Adenovirus And Bovine Coronavirus., Kevin Lane

Theses

This investigation involved the analysis of selected gastroenteritis viruses, adenovirus and bovine coronavirus, obtained from faecal samples, collected from hospital and diagnostic laboratories respectively in Cork city, Ireland. The viruses isolated from these samples were subjected to analysis using cell culture and nucleic acid extraction. In addition adenovirus was analysed using PCR and PCR followed by restriction endonuclease digest. Many samples contained more than one gastroenteritis virus, the virus of interest and a natural co-infection with rotavirus species. Rotavirus infections in Ireland have been extensively studied and we were interested in studying singly and doubly infect samples.

Bovine enteric coronavirus …

The Analysis And Characterisation Of Rotavirus And Norovirus Circulating In The South Of Ireland, Norma Reidy

Theses

On average more than 3,000 cases of rotavirus diarrhea are detected annually amongst Irish children less than 3 years of age. Rotavirus disease occurs mainly during the winter months, with peak infections detected from February to April. The symptoms of disease are diarrhea and dehydration, with 80% of patients requiring hospitalisation, The mean duration of hospitalisation is 3-4 days, with an average cost per day of €409. Eighteen percent of infections were detected in infants less than 6 months of age, suggesting inadequate maternal derived protection from existing strains, or the existence of novel strains in Ireland with little or …

The Investigation Of Rna Viruses Using The Atomic Force Microscope, Michelle Moloney

Theses

The subject of viral entry into, and subsequent progeny virus egress, from cultured cells, has been extensively studied using numerous scientific techniques in fields ranging from cell culture to biochemistry to microscopy. The atomic force microscope (AFM) is a novel, developing instrument with unprecedented capabilities and this study was undertaken to observe enveloped Semliki Forest virus (SFV) and non-enveloped Theiler's Murine Encephalomyelitis virus (TMEV) entry into and egress from cultured cells using the AFM. Virus particle structure of both enveloped and non-enveloped viruses was also directly examined.

Cytopathic effect studies revealed that TMEV's are unable to produce progeny viruses in …

The Molecular Epidemiology Of Rotavirus In Ireland, Fiona O'Halloran

Theses

Between 1997 and 1998, 3,136 cases of rotavirus diarrhoea were detected in Irish children less than 2 years of age. Hospital inpatients accounted for 80% of these infections, with the remainder being diagnosed in general practice. A large percentage of infections were detected in neonates, suggesting a possible inadequate maternal derived protection from existing indigenous strains. This feature suggested the possible existence of ‘novel’ strains circulating in Ireland. No data describing the epidemiology of rotavirus strains in this country currently exists. Furthermore no assessment of the potential health-economic impact or quantitation of potential disease burden was ever undertaken

Three hundred …