Pharmaceutical Microbiology (PHRM361) - University of Kwazulu-Natal PDF

PHARMACEUTICAL MICROBIOLOGY (PHRM361) Dr Mbuso Faya, PhD. Discipline of Pharmaceutical Sciences, UKZN, South Africa PHARMACEUTICAL MICROBIOLOGY Examine and understand the concepts of Quality Control and Quality Assurance as they apply to the pharmaceutical industry....

PHARMACEUTICAL MICROBIOLOGY (PHRM361) Dr Mbuso Faya, PhD. Discipline of Pharmaceutical Sciences, UKZN, South Africa PHARMACEUTICAL MICROBIOLOGY Examine and understand the concepts of Quality Control and Quality Assurance as they apply to the pharmaceutical industry. Acquire knowledge of microbial production and assay methods. Perform laboratory procedures while practising safe, aseptic laboratory technique. Enumerate and identify micro-organisms from commercial products. Evaluate the antimicrobial effectiveness of disinfectants, preservatives and antibiotics. Analyze the effectiveness of antibiotics (Second Term). Perform microbial assays  Introduction Microbiology is the — ‘scientific study of the microorganisms’. microorganism invariably refers to the minute living body not perceptible to the naked eyes, especially a bacterium or viruses. Importantly, microorganisms may be carried from one host to another as follows (a) Animal Sources. Certain organisms are pathogenic for humans as well as animals and may be communicated to humans via direct, indirect, or intermediary animal hosts. (b) Airborne. Pathogenic microorganisms in the respiratory tract may be discharged from the mouth or nose into the air and usually settle on food, dishes or clothing. They may carry infection if they resist drying. (c) Contact Infections. Direct transmission of bacteria from one host to another viz., sexually transmitted diseases (STD). (d) Foodborne. Food as well as water may contain pathogenic organisms usually acquired from the handling the food by infected persons or via faecal or insect contamination.  Introduction (e) Fomites. Inanimate objects e.g., books, cooking utensils, clothing or linens that can harbour microorganisms and could serve to transport them from one location to another. (f) Human Carriers. Persons who have recovered from an infectious disease do remain carriers of the organism causing the infection and may transfer the organism to another host. (g) Insects. Insects may be the physical carriers, for instance housefly (Musca domestica), or act as intermediate hosts, such as the Anopheles mosquito. (h) Soilborne. Spore-forming organisms in the soil may enter the body via a cut or wound. Invariably fruits and vegetables, particularly root and tuber crops, need thorough cleansing before being eaten raw. Introduction Microbiology is the specific branch of ‘biology’ that essentially deals with the elaborated investigation of ‘microscopic organisms’ termed as microbes, that are composed of only one cell. These are typically either unicellular or multicellular microscopic organisms that are distributed abundantly both in the living bodies of plants and animals and also in the air, water, soil, and marine kingdom. CLASSIFICATION OF MICRO- ORGANISMS CLASSIFICATION OF MICRO- ORGANISMS Taxonomy Organizing, classifying and naming living things Identifying and classifying organisms according to specific criteria Each organism placed into a classification system Domain Kingdom Phylum Class 1. Find out what these Order mean 2. Discuss the Family significance of Genus taxonomy species 3. Name one tool used in the lab as a classification tool CLASSIFICATION OF MICRO- ORGANISMS 3 Domains 1. Discuss the Eubacteria importance of peptidoglycan true bacteria, peptidoglycan2. Drugs used to Archaea target it odd bacteria that live in extreme environments, high salt, heat, etc. (usually called extremophiles) Eukarya have a nucleus & organelles (protists, fungi, algae) CLASSIFICATION OF MICRO- ORGANISMS  CLASSIFICATION OF MICRO- ORGANISMS Eukaryotes nucleus nuclear membrane cytoplasm with self- replicating organelles  CLASSIFICATION OF MICRO- ORGANISMS Prokaryotes ( Bacteria, rickettsia, chlamydia) Bacteria No true nucleus Lack internal membranes Rigid cell wall Reproduce by binary fission Contain both DNA and RNA Metabolic enzymes Visible under light microscope Cannot pass through a 0.2micrometer membrane.  CLASSIFICATION OF MICRO- ORGANISMS Akaryocytes Cells without a nucleus Name derived from the Greek prefix "a-", meaning "without" and the Greek "karyo-", meaning "nut" or "kernel". Akaryocytes also include viruses since they lack a nucleus but have instead, a central core of RNA or DNA. Akaryocytes are not part of the seven main ranks of taxa. 1.Capsid encapsulates the viral genome 2. protection, attachment, stability 3. Entry into cells 4. Release and packaging of  CLASSIFICATION OF MICRO- ORGANISMS Viruses They have no metabolic system of their own. They depend upon the machinery of the host cell for replication (obligate intracellular parasites). They have either DNA or RNA genomes, but lack ribosomes and other factors needed for translation. Thus, they are dependent on the host cells for production of viral proteins. Their genomes encode minimal information to ensure the following: 1) genome replication and packaging; 2) production of viral proteins; and 3) subvert cellular functions to allow the production of virions. Some viruses (bacteriophages) infect prokaryotic cells, while others infect eukaryotic cells. Some viruses destroy cells, producing disease; other persist in infected cells either in a latent or persistent state; and other may cause cellular malignant transformation. Structure and function of microorganisms Structure and function of microorganisms Prokaryotic Cells Eukaryotic Cells Small cels (< 5um) Larger cells (>10um) Always unicellular Multicellular No nucleus or any membrane Defined nucleus and other bound organelles, e.g. membrane bound organelles mitochondria DNA is circular, without DNA is linear – associated with proteins proteins to form Chromatin Small Robosomes (70S) Large Ribosomes (80S) cytoskeleton Has a cytoskeleton Motiliyt (flagellum) – rigid Motility by flexible waving cilia motility or flagellae (made of tubulin) Cell division – binary fission Cell division by mitosis or meiosis Reproduction – asexual Reproduction is asexual or sexual Huge variety of metabolic Common metabolic pathways pathways Structure and function of microorganisms 3. Basic structural forms of viruses Viruses are minimally composed of a nucleic acid genome (DNA or RNA) and a protein coat. Many viruses contain an external membrane called an envelope. Structure and function of microorganisms Icosahedral symmetry The genetic material is fully enclosed inside of the capsid. Viruses with icosahedral structures are released into the environment when the cell dies, breaks down and lyses, thus releasing the virions. poliovirus, rhinovirus, and adenovirus possess an icosahedral symmetry Structure and function of microorganisms Envelope helical structure that is surrounded by a lipid bilayer membrane, meaning the virus is encased or enveloped. The envelope of the virus is formed when the virus is exiting the cell via budding, and the infectivity of these viruses is mostly dependent on the envelope Examples: influenza virus, Hepatitis C and HIV. Structure and function of microorganisms Complex combination of icosahedral and helical shape and may have a complex outer wall or head-tail morphology The head-tail morphology structure is unique to viruses that only infect bacteria and are known as bacteriophages. Example: Bacteriophage, Poxvirus Structure and function of microorganisms Helical capsid with a central cavity or hollow tube that is made by proteins arranged in a circular fashion, creating a disc like shape. All filamentous viruses are helical in shape. Example: tobacco mosaic virus. Structure and function of microorganisms Viral Entry A virus receptor can be defined as a host cell surface component recognized by the virus as a gateway to entry into the cell Ideally, a virus receptor would fulfill three main characteristics: (1) a physical interaction between the virus and the receptor should be demonstrated; (2) occupying the virus-binding site of the receptor (e.g., with an antibody directed against the receptor, should inhibit virus infection); and (3) the cellular sensitivity to virus infection should correlate with receptor expression.  Structure and function of microorganisms Viral receptors The CD4 transmembrane protein expressed by a subset of human T-lymphocytes is widely considered as the main receptor for HIV-1. Cellular protein TIM-1 (-cell immunoglobulin and mucin domain 1 ) acts as a receptor for Ebola virus and Marburg virus Domain structures of TIM-1 and TIM-3 Life cycles of microorganisms Life cycles of microorganisms Viral Life cycle Bacterial Life cycle Ecological Relationships Microbial interactions Host-parasite interactions The environment Microbial interactions The triad consists of an external agent, a susceptible host, and an environment that brings the host and agent together. In this model, disease results from the interaction between the agent and the susceptible host in an environment that supports transmission of the agent from a source to that host ACQUIRING INFECTIOUS AGENTS PORTAL OF ENTRY/EXIT INGESTION INHALATION DIRECT PENETRATION Trauma or Surgical Procedure Needlestick Arthropod Bite Sexual Transmission Transplacental  ACQUIRING INFECTIOUS AGENTS (cont.) COLONIZATION: the successful occupation of a new habitat by a species not normally found in this niche Adherence (attachment): close association of bacterial cells and host cells generally characterized by receptors on target sites Adhesin: structure or macromolecule located on the surface of a cell or extracellularly that facilitates adherence of a cell to a surface or to another cell; site of attachment is often a specific receptor and host cell receptors are often sugar moieties (lectin), but the adherence may also be nonspecific  ACQUIRING INFECTIOUS AGENTS (cont.) INVASION: the entry and spread throughout the cells and/or tissues of the host; specific recognition of receptor sites on target cells enhances pathogenic advantage Invasins (invasive factors): structures or macromolecules that facilitate invasion by a pathogenic microorganism MULTIPLICATION: the ability of a microorganism to reproduce during an infection; influenced by underlying disease, immunologic status, antibiotic treatment, nutrient availability  TRANSMISSION OF DISEASE ENTRANCE, COLONIZATION, PENETRATION: Dependent upon Age, Sex, Nutrition, Immunologic State and General Health of Host, and Bacterial Virulence Factors VECTOR: a carrier, especially the animal that transfers an infectious agent from one host to another, usually an ARTHROPOD CARRIER (Carrier State): symptomless individual who is host to a pathogenic microorganim with the potential to pass the pathogen to others NOSOCOMIAL INFECTIONS: an infection acquired in a hospital setting that was not present in the host prior to admission, generally occurring within 72 hours of admission  PATHOGENICITY vs. VIRULENCE PATHOGENICITY: the quality of producing disease or the ability to produce pathologic changes or disease VIRULENCE: a measure of pathogenicity; a measurement of the degree of disease-producing ability of a microorganism as indicated by the severity of the disease produced; commonly ascertained by measuring the dosage required to caused a specific degree of pathogenicity one general standard is the LD50 (lethal dose 50%) Bacterial Virulence Mechanisms Adherence (Colonization) Invasion Degradative enzymes Exotoxins Endotoxin Induction of excess inflammation Evasion of phagocytic & immune clearance Byproducts of growth (gas, acid) Superantigen Resistance to antibiotics  INFECTION vs. DISEASE INFECTION: the colonization and/or invasion and multiplication of pathogenic microrganisms in the host with or without the manifestation of disease DISEASE: an abnormal condition of body function(s) or structure that is considered to be harmful to the affected individual (host); any deviation from or interruption of the normal structure or function of any part, organ, or system of the body  INFECTION vs. DISEASE (Definitons) BENIGN: a non-life or non-health threating condition MALIGNANT: a disease tending to become progressively worse (MORBIDITY = illness) and potentially result in death (MORTALITY = death) CONTAGIOUS: capable of being transmitted from one host to another; communicable; infectious INFECTIOUS DOSE: number of pathogenic organisms required to cause disease in a given host  KOCH'S POSTULATES Four criteria that were established by Robert Koch to identify the causative agent of a particular disease, these include: 1. the microorganism (pathogen) must be present in all cases of the disease 2. the pathogen can be isolated from the diseased host and grown in pure culture 3. the pathogen from the pure culture must cause the same disease when inoculated into a healthy, susceptible laboratory animal 4. the pathogen must be reisolated from the new host and shown to be the same as the originally inoculated pathogen KOCH'S POSTULATES Pathogenicity The development of novel drugs, vaccines and other therapeutics going into the future will be highly dependent on the knowledge gained from investigating host-pathogen interactions.  Pharmaceutical Microbiology concerned with the use of microorganisms in pharmaceutical development maintaining contamination control. Health Care Infection control Possible contaminants of pharmaceutical products Good manufacturing practice