Lecture3 Diagnosis and Control of Infection PDF

Diagnosis and control of infection Dr. Monica Agromayor Department of Infectious Diseases Faculty of Life Sciences and Medicine Principles of disease transmission Epidemiology Science that studies when and where diseases occur and how they are transmitted in a population. Epidemiologists collect...

Diagnosis and control of infection Dr. Monica Agromayor Department of Infectious Diseases Faculty of Life Sciences and Medicine Principles of disease transmission Epidemiology Science that studies when and where diseases occur and how they are transmitted in a population. Epidemiologists collect information to determine: Aetiology: identification of the pathogen causing the disease Predisposing factors: age, sex, lifestyle, etc. – identify susceptible populations Incidence: number of individuals acquiring disease in a given time period. Prevalence: number of individuals with disease in a given time period. Mode of transmission Public health policy and prevention Disease development and transmission Disease development and transmission Chain of infection is a concept used to explain how a patient can acquire an infection from another person It is used by epidemiologists and clinical microbiologists to develop strategies to prevent and control epidemics. The components of the chain will vary depending on the microbe and the disease it causes. Transmission of any given infection is stopped by breaking one or more of the links. Disease development and transmission The spread of infection is the final requirement for a successful pathogen. Two main factors affect the spread of infection: Reservoirs of infectious organisms - places where pathogens can grow and accumulate Modes of transmission – the various ways in which pathogens move from place to place Reservoirs Habitat in which an infectious agent normally lives, grows, RESEVOIR and multiplies. Usually source from which it is transmitted to a susceptible host. Zoonotic diseases spread NON HUMAN ANIMAL between animals and people. LIVING The natural reservoir of some diseases remains unknown. CASE CARRIER Reservoirs Habitat in which an infectious agent normally lives, grows, RESEVOIR and multiplies. Usually source from which it is transmitted to a susceptible host. Zoonotic diseases spread NON HUMAN ANIMAL between animals and people. LIVING The natural reservoir of some diseases remains unknown. CASE CARRIER Reservoirs Habitat in which an infectious agent normally lives, grows, RESEVOIR and multiplies. Usually source from which it is transmitted to a susceptible host. Zoonotic diseases spread NON HUMAN ANIMAL between animals and people. LIVING The natural reservoir of some diseases remains unknown. CASE CARRIER Reservoirs Habitat in which an infectious agent normally lives, grows, RESEVOIR and multiplies. Usually source from which it is transmitted to a susceptible host. Zoonotic diseases spread NON HUMAN ANIMAL between animals and people. LIVING The natural reservoir of some diseases remains unknown. CASE CARRIER Emerging disease Unrecognized infection, or a previously recognized infection that has expanded into a new ecological niche, often accompanied by a significant change in pathogenicity. Many emerging diseases are zoonotic - an animal reservoir incubates the organism, with only occasional transmission into human populations. Emerging disease Unrecognized infection, or a previously recognized infection that has expanded into a new ecological niche, often accompanied by a significant change in pathogenicity. Many emerging diseases are zoonotic - an animal reservoir incubates the organism, with only occasional transmission into human populations. Can be caused by: Newly identified species (e.g. HIV and AIDS) Newly identified strains that have evolved from a known infection (e.g. influenza) Ecological changes that alter the composition and size of reservoirs (e.g. Lyme disease) Spread to a population in a new area of the globe (e.g West Nile fever) Re-emerging infections like drug resistant tuberculosis Nosocomial (hospital-acquired) infections, such as Methicillin-resistant S. aureus Emerging disease Disease transmission Main modes of transmission: Contact transmission Indirect transmission – vehicle or vector Horizontal (vs vertical) Contact transmission A healthy person is exposed to pathogens by either touching or being close to an infected person or object. Direct Contact Transmission: Person- to-person transmission (touching, kissing, sexual intercourse). No intermediate object is involved Examples: Hepatitis A, Smallpox, Staphylococcal infections, mononucleosis, sexually transmitted diseases such as syphilis or HIV/AIDS. Contact transmission A healthy person is exposed to pathogens by either touching or being close to an infected person or object. Indirect Contact Transmission: The microbe is transferred via a nonliving object or fomite, such as towels, eating utensils, thermometers, stethoscopes, bedding, clothes, money, and needles. Examples: Herpes simplex virus, Cytomegalovirus, Giardia, Impetigo Contact transmission A healthy person is exposed to pathogens by either touching or being close to an infected person or object. Droplet Transmission: Microbes are spread in mucus droplets that travel short distances (less than 1 meter). It can occur through sneezing, coughing, or talking. Typical of respiratory viruses (e.g., influenza, adenovirus, respiratory syncytial virus, human metapneumovirus), Bordetella pertussis, Pneumococci, Diphtheria and Rubella. Vehicle transmission Transmission of disease via medium such as water, food, air, blood, body fluids, and intravenous fluids. Waterborne Transmission: Usually caused by water contaminated with sewage. Airborne Transmission: Not to be confused with droplet transmission, is due to inhalation of small pathogens and particles (e.g. bacterial and fungal spores) that are suspended in air and can travel long distances. Foodborne Transmission: Typically due to bad sanitation practices leading to contamination of food with pathogens Examples: Anthrax, tuberculosis, salmonella, cholera, typhoid, legionella Vector transmission Transmission of disease via animals that carry disease from one host to another. Insects are most important animal vectors. Mechanical Transmission: Passive transport of pathogens on vector’s body. Flies are the most common vector. Most of the diseases can also be contracted more directly through contaminated food, water, air, hands and person-to-person contact Examples include enteric infections (dysentery, diarrhoea, typhoid or cholera) and eye infections (trachoma and conjunctivitis) Vector transmission Transmission of disease via animals that carry disease from one host to another. Insects are most important animal vectors. Biological Transmission: Pathogen spends part of its life cycle in the vector and transmission to the host is through a bite. Examples include malaria, Zika virus, Dengue fever, schistosomiasis and rabies Horizontal vs vertical transmission Transmission from mother to child is called vertical transmission and can occur in utero across placenta, at the time of delivery or during breast feeding. Person-to-person transmission that is not between mother and offspring is called horizontal transmission. Examples include HIV, Rubella and toxoplasmosis Detection and diagnosis Koch’s postulates Diseased animal Healthy animal.. 1. The microorganism must be found in abundance in all organisms suffering from. 1. Observe sample.... under microscope the disease, but not in healthy organisms. Suspected pathogen Red blood cells 2. The microorganism must be isolated from a diseased organism and grow in pure No pathogen culture. 2. Culture sample from diseased or present healthy animals 3. The cultured microorganism should cause Cultured pathogen disease when introduced into a healthy 3. Inoculate healthy animal with suspected pathogen organism. Diseased animal 4. The microorganism must be reisolated from the inoculated, diseased experimental host and identified as being identical to the 4. Culture original specific causative agent. Koch’s postulates today Koch’s principles do not apply to all diseases: Exceptions to the first postulate: Asymptomatic or subclinical infection carriers are a common feature of many infectious diseases e.g. cholera or typhoid fever and viral infections such as polio, herpes simplex, HIV and hepatitis C Exceptions to the second postulate: Some microbes cannot be grown in vitro or there are no susceptible animal species e.g. Treponema pallidum (syphillis), Mycobacterium leprae (leprosy) and wart viruses Exceptions to the third postulate: not all organisms exposed to an infectious agent will acquire the infection e.g. resistance to malaria conferred by possessing at least one sickle cell allele. Koch’s postulates continue to inform the approach to microbiologic diagnosis but fulfillment of all four postulates is no longer required to demonstrate causality Laboratory diagnosis Two main methods are used in a diagnostic laboratory to confirm infection: Direct detection methods – clinical specimen is examined for the presence of a microbe or its products. These include culture, microscopy and molecular methods Indirect detection (serological) methods – blood and other body fluids are examined for the presence of antibodies against a pathogen The specimen to be collected from the patient needs to be selected taking into account the pathogenesis of the infection. Direct methods: culture of bacteria Method used to propagate microorganisms by allowing them to grow in predetermined culture media under controlled laboratory conditions (e.g. temperature, air supply, light, pH). Depends on culture of microorganism mainly on solid nutrient media (agar plates) to produce colonies. A colony is composed of thousands of bacteria growing on the surface that originate from a single cell. Direct methods: culture of bacteria Method used to propagate microorganisms by allowing them to grow in predetermined culture media under controlled laboratory conditions (e.g. temperature, air supply, light, pH). Depends on culture of microorganism mainly on solid nutrient media (agar plates) to produce colonies. A colony is composed of thousands of bacteria growing on the surface that originate from a single cell. Types of medium: Defined medium: if the exact chemical composition is known. Direct methods: culture of bacteria Method used to propagate microorganisms by allowing them to grow in predetermined culture media under controlled laboratory conditions (e.g. temperature, air supply, light, pH). Depends on culture of microorganism mainly on solid nutrient media (agar plates) to produce colonies. A colony is composed of thousands of bacteria growing on the surface that originate from a single cell. Types of medium: Defined medium: if the exact chemical composition is known. Enrichment medium: contains some component that permits the growth of specific types or species of bacteria, usually because they alone can utilize the component from their environment. Selective medium: Culture media designed to support the growth of only specific microorganisms (e.g. selection done by adding antibiotics or lacking amino acids). Direct methods: culture of bacteria Method used to propagate microorganisms by allowing them to grow in predetermined culture media under controlled laboratory conditions (e.g. temperature, air supply, light, pH). Depends on culture of microorganism mainly on solid nutrient media (agar plates) to produce colonies. A colony is composed of thousands of bacteria growing on the surface that originate from a single cell. Types of medium: Defined medium: if the exact chemical composition is known. Enrichment medium: contains some component that permits the growth of specific types or species of bacteria, usually because they alone can utilize the component from their environment. Selective medium: Culture media designed to support the growth of only specific microorganisms (e.g. selection done by adding antibiotics or lacking amino acids). Differential medium: Distinguishes closely related microorganism growing on the same media on a difference in the colony appearance due to the presence of certain dyes or chemicals in the media. Direct methods: culture of viruses Propagation requires cell cultures as viruses only replicate in living cells. Cells that support viral replication are called permissive. Multiplicity of infection (MOI) refers to the number of virions that are added per cell during infection. A virus-infected cell can show dramatic changes in appearance or cytopathic effects (CPE) such as cell rounding and detachment from surface, cell fusion (syncytia) or inclusion bodies. Direct methods: culture of viruses Propagation requires cell cultures as viruses only replicate in living cells. Cells that support viral replication are called permissive. Multiplicity of infection (MOI) refers to the number of virions that are added per cell during infection. A virus-infected cell can show dramatic changes in appearance or cytopathic effects (CPE) such as cell rounding and detachment from surface, cell fusion (syncytia) or inclusion bodies. A B C Monolayer of healthy cells Infected cells showing CPE Direct methods: microscopy Microscopy and molecular techniques based on detection of nucleic acids provide a rapid indication of microbial infection in a matter of hours. Used either with wet or fixed samples Can be used in combination with stains. Can study bacteria morphology and staining characteristics. Used to determine virus-induced CPE. To see virus morphology electron microscopy is needed. Direct methods: microscopy Microscopy and molecular techniques based on detection of nucleic acids provide a rapid indication of microbial infection in a matter of hours. Used either with wet or fixed samples Can be used in combination with stains. Can study bacteria morphology and staining characteristics. Used to determine virus-induced CPE. To see virus morphology electron microscopy is needed. Direct methods: microscopy Microscopy and molecular techniques based on detection of nucleic acids provide a rapid indication of microbial infection in a matter of hours. Used either with wet or fixed samples Can be used in combination with stains. Can study bacteria morphology and staining characteristics. Used to determine virus-induced CPE. To see virus morphology electron microscopy is needed. Direct methods: detection of nucleic acids Molecular tests can detect the presence of bacterial and viral DNA or RNA in a patient specimen. Labelled probe Sample Can be used to detect organisms that are slow or difficult to grow in the laboratory, antibiotic resistance genes or virulence factors. Very sensitive and specific. Two main types: Hybridisation techniques with nucleic acid probes Fully complementary strands bind strongly Direct methods: detection of nucleic acids Molecular tests can detect the presence of bacterial and viral DNA or RNA in a patient specimen. Labelled probe Sample Can be used to detect organisms that are slow or difficult to grow in the laboratory, antibiotic resistance genes or virulence factors. Very sensitive and specific. Two main types: Hybridisation techniques with nucleic acid probes Fully complementary strands bind strongly Direct methods: detection of nucleic acids Molecular tests can detect the presence of bacterial and viral DNA or RNA in a patient specimen. Can be used to detect organisms that are slow or difficult to grow in the laboratory, antibiotic resistance genes or virulence factors. Very sensitive and specific. Two main types: Hybridisation techniques with Original DNA to be replicated nucleic acid probes (template) Polymerase chain reaction (PCR) dNTPs Primer Indirect methods: serological tests Determine the presence of antibodies in serum or microbial antigens in tissue or body fluids. Titre refers to antibody concentration in the sample and is associated with the number of times one can dilute a sample and still detect the antibody. Indirect methods: serological tests Determine the presence of antibodies in serum or microbial antigens in tissue or body fluids. Titre refers to antibody concentration in the sample and is associated with the number of times one can dilute a sample and still detect the antibody. Paired sera samples need to be collected during acute phase (5-7 days after onset of symptoms) and convalescence phase of infection (after 2-4 weeks). At least a four-fold rise in specific antibody titre between acute and convalescent samples (seroconversion) must be found for a diagnosis to be made. Presence of IgM but not IgG antibodies can be an indication of current active infection. Do not distinguish between previous or current infection. Serological methods are retrospective. Indirect methods: serological tests Determine the presence of antibodies in serum or microbial antigens in tissue or body fluids. Titre refers to antibody concentration in the sample and is associated with the number of times one can dilute a sample and still detect the antibody. Paired sera samples need to be collected during acute phase (5-7 days after onset of symptoms) and convalescence phase of infection (after 2-4 weeks). At least a four-fold rise in specific antibody titre between acute and convalescent samples (seroconversion) must be found for a diagnosis to be made. Presence of IgM but not IgG antibodies can be an indication of current active infection. Do not distinguish between previous or current infection. Serological methods are retrospective.  Direct vs indirect methods Method Advantages Disadvantages Culture Confirms presence of organism Can take long Organism is multiplied and can be used Organism need to be capable of growing for additional testing in vitro Microscopy Quick and easy preliminary results Not very specific Detection of nucleic acids Results available within hours Previous knowledge of DNA sequence Very sensitive and specific needed Serological tests Not limited to blood serum (other body Long length of time required to obtain fluids also suitable) paired sera Well established, inexpensive and easy Specific antibodies not always available to perform Not suitable for agents that produce Can be used for multiple sample clinical disease before the appearance of analysis at a time antibodies Some conditions may not produce detectable antibodies Prone to false positive results due to antigenic cross-reactivity between related agents Strategies to treat infection Selective toxicity Antimicrobial agents need to inhibit the growth of the Penicillium colony microbe while doing minimal damage to the patient. Achieved by exploiting the differences between the metabolism and structure of microorganisms and the human cells they infect. Compared to the number of drugs available to treat bacterial infection, the number of antiviral drugs is very small viral replication is intimately linked to normal cellular functions so selective toxicity is difficult to obtain. Antimicrobial agents can be natural products (e.g. antibiotics from fungal sources) or synthetic if they are chemically designed in the lab. Antibacterial agents Type of antimicrobial drug used in the treatment and prevention of bacterial infections. They may either kill (bactericidal) or inhibit the growth of bacteria (bacteriostatic) May be classified as either broad spectrum or narrow spectrum depending on how many types of microorganism are naturally susceptible to their action Also classified depending on their chemical structure and site of action: Molecules that inhibit cell wall synthesis Molecules that inhibit the function of the bacterial plasma membrane Molecules that inhibit the synthesis of nucleic acids Molecules that inhibit the synthesis of proteins Antibacterial agents Major targets of common antibacterial agents Principle of action Cell wall synthesis The cell wall is rich in peptidoglycans, a compound unique to bacteria. Plasma membrane Injure plasma membrane by disrupting function membrane potential. Some target specifically the LPS within Gram-negative outer membranes. Nucleic acid Block bacterial enzymes or metabolic synthesis pathways that produce essential precursors needed for DNA and mRNA synthesis Protein synthesis Exploit the differences between eukaryotic and prokaryotic ribosomal proteins, RNAs and associated enzymes Antiviral agents Major steps targeted by common antiviral agents Principle of action Attachment and entry Inhibit fusion of viral envelope or attachment to receptor Nucleic acid Molecules that target viral DNA and synthesis RNA polymerases Assembly and budding Inhibit viral proteins needed for virion maturation and/or release Antimicrobial drug resistance Antimicrobial resistance is the ability of a microorganism to survive and multiply in the presence of an antimicrobial agent that would normally inhibit or kill this particular kind of organism. Bacterial resistance strategies: By preventing drug from reaching its target by reducing its ability to penetrate the cell By inactivation of drug via modification or degradation By expulsion of the drug from the cell via general or specific efflux pumps By modification of the drug’s target site within the bacteria Antimicrobial drug resistance Antimicrobial resistance is the ability of a microorganism to survive and multiply in the presence of an antimicrobial agent that would normally inhibit or kill this particular kind of organism. Viral resistance strategies: Results from spontaneous mutations in the viral genome during viral replication Mutations are within the target of the antiviral drug The error-prone polymerase enzyme in RNA viruses cause these viruses to develop resistance more frequently than DNA viruses Special concern during extended therapy for chronic infections (e.g. HIV, HBV and HCV) Combination therapy with more than one agent is commonly used to delay appearance of resistance Prevention of disease transmission Immunological memory First exposure Clonal Memory cells are long- Second Stronger and more rapid to antigen expansion lived, continue to exposure to response reproduce antigen Effector cells carry out immediate response; short lived Primary response Secondary response  Types of adaptive immunity ADAPTIVE IMMUNITY ACTIVE PASSIVE NATURAL ARTIFICIAL NATURAL ARTIFICIAL Exposure to infectious agent Vaccination Maternal antibodies Injected Antibodies Artificial passive immunisation Immunity is transferred by administration of antibodies to a non immune individual. Used when there is no time to wait for the development of active immunity, or when no effective active vaccine exists. The antibodies used can be either of human origin or produced in animals Artificial active immunisation Immunity develops after the immune system encounters an antigen and is the basis for VACCINATION. Thus, vaccination is exposing a person to material that is antigenic but NOT pathogenic, inducing adaptive immunity and memory. At the population level, the success of a vaccination programme depends on herd immunity. Herd immunity: in contagious diseases that are transmitted from individual to individual, chains of infection are likely to be disrupted when large numbers of a population are immune or less susceptible to the disease. The proportion of the population required to be immune to reach herd immunity depends on a number of factors and it’s different for each disease. Requirements of an effective vaccine Safe, with no or few side effects – it must not cause disease!! Give long lasting, appropriate protection against the natural form of the pathogen Stimulate both a humoral and cell-mediated immune response and the production of memory cells. Low in cost Stable with long shelf life and no special storage requirements Easy to administer