Prelims Micro Lec.pdf

I. History of Microbiology Nucleus No nuclear Classic membrane membrane- Anton van Leeuwenhoek: Known as the...

I. History of Microbiology Nucleus No nuclear Classic membrane membrane- Anton van Leeuwenhoek: Known as the bound nucleus “father of protozoology and bacteriology”. DNA Circular, Linear, found located in in nucleus o Discovered microorganisms nucleoid (referred to as "wee beasties" or Membrane- Absent Present (ER, "animalcules") using a homemade bound mitochondria, microscope. Organelles etc.) o His discoveries led to the Ribosomes 70S ribosomes 80S ribosomes (50S + 30S (60S + 40S recognition of the existence of subunits) subunits) microbes, many of which do not Cell Wall Peptidoglycan Cellulose cause disease. (bacteria) (plants), chitin (fungi) II. Bacteria: Overview and Structure Flagella Simple, rotary Complex, action coordinated Bacteria are Prokaryotes: sliding MTs o Unicellular organisms lacking a true Electron Cell membrane Mitochondria nucleus (i.e., no nuclear Transport inner membrane). membrane o They lack membrane-bound Pili/Fimbriae Present Absent organelles such as mitochondria, endoplasmic reticulum (ER), and Golgi bodies. IV. Unique Structures in Bacteria Prokaryote vs. Eukaryote: Major structural 1. Plasmids: and functional differences: o Size: Prokaryotes are much smaller o Small, circular DNA molecules. (0.4–2 µm in diameter) compared to eukaryotes (10–100 µm). o Often found in Gram-negative o Nucleus: Prokaryotes have no true bacteria. nucleus; DNA is in a nucleoid region. Eukaryotes have a o Carry genes that provide antibiotic membrane-bound nucleus. resistance and virulence factors. o DNA structure: ▪ Prokaryotes: Circular DNA, 2. Cell Wall Composition: complexed with RNA; o Bacterial cell walls are composed of plasmids often present. peptidoglycan (absent in ▪ Eukaryotes: Linear DNA eukaryotes). complexed with histones. o Reproduction: Prokaryotes o Eukaryotic cell walls, when present, reproduce asexually via binary contain cellulose, chitin, or other fission. Eukaryotes can reproduce glycans. both sexually and asexually. 3. Flagella: II. Prokaryotic vs. Eukaryotic Cell Characteristics o Bacterial flagella are simpler, (Table 1.1 comparison points) composed of flagellin proteins and powered by rotary motion. Characteristic Prokaryote Eukaryote Size 0.4–2 µm 10–100 µm diameter diameter  o Eukaryotic flagella are more o Yeasts: Unicellular fungi that complex, made of microtubules reproduce asexually. with coordinated sliding action. ▪ "True" yeasts do not form hyphae or mycelia. 4. Ribosomes: o Most fungi: Multicellular, with reproduction occurring both o Bacteria have 70S ribosomes sexually and asexually. (distinct from the 80S ribosomes in ▪ Hyphae: Filaments that eukaryotes). interweave to form mycelia. o Ribosomes are the site of protein ▪ Molds: Filamentous fungi synthesis. that reproduce similarly. ▪ Dimorphic fungi: Can grow V. Reproduction as yeasts at 37°C (human body temperature) and as Prokaryotes reproduce asexually by binary filamentous forms at 22°C fission. (room temperature). Eukaryotes can reproduce either sexually ▪ Some systemic fungal (meiosis) or asexually (mitosis). infections are caused by dimorphic fungi. Key Differences to Remember for Exams Viruses: 1. Prokaryotes vs. Eukaryotes: Size, organelles, and DNA structure are major differentiators. Smallest infectious particles, not visible 2. Bacterial Structures: Understand the roles of under a standard microscope. plasmids, cell wall, flagella, and ribosomes. o Can be identified by their effects on 3. Bacterial Reproduction: Binary fission infected cells (e.g., syncytium (asexual) is the main method for formation, rounding up of cells). prokaryotes. o Characteristics of viruses: 4. Energy Production: Electron transport ▪ Consist of either DNA or happens in the cell membrane in bacteria RNA (never both). (no mitochondria present). ▪ Acellular: Lacking cytoplasmic membranes, Parasites: surrounded by a protein coat. Eukaryotic parasites can be unicellular or ▪ Obligate intracellular multicellular. parasites: Require host o Protozoa: Unicellular organisms cells for replication and from the kingdom Protista that use the host's machinery obtain nutrition via ingestion. for reproduction. ▪ They are classified by their ▪ Host specificity: Viruses locomotion: flagella (whip- typically infect specific like), pseudopodia (false host cells (e.g., HIV targets feet), or cilia (eyelash-like). T-helper cells). o Multicellular parasites: Examples ▪ Bacteriophage: A virus include tapeworms, which can that infects bacteria. reach lengths of 7–10 meters. Classification/Taxonomy: Fungi: Taxonomy: Classification of organisms into Fungi are heterotrophic eukaryotes that groups called taxa. absorb nutrients.  o Based on similarities in genotype Taxonomy: Be familiar with classification (genetic makeup) and phenotype terms and the distinctions between (observable traits). genotype and phenotype. o Taxa include domain, kingdom, Domains: Know the three domains— phylum, class, order, family, genus, Bacteria, Archaea, and Eukarya—and the species, and subspecies. unique features of archaea. Bacterial species: Often divided into serotypes, biotypes, or subspecies based on Prokaryotic Cell Structure minor differences. Nomenclature: Follows specific rules: 1. Cytoplasmic Structures: o Family names end in "-aceae" (e.g., Micrococcaceae). o Nucleus: Prokaryotes lack a o Genus and species names are membrane-bound nucleus; their italicized or underlined (e.g., genetic material is a single circular Staphylococcus aureus). chromosome located in the nucleoid. Classification by Cellular Type: o Ribosomes: Prokaryotic ribosomes are 70S in size (composed of 50S Three domains of life: Bacteria, Archaea, and 30S subunits). and Eukarya. o Archaea: Similar to prokaryotes but o Storage Granules: Prokaryotes may more closely related to eukaryotic contain cytoplasmic granules for cells. Often found in extreme storage of polysaccharides, lipids, environments (e.g., halophiles and or polyphosphates. thermophiles). o Prokaryotes: Include bacteria and o Endospores: Certain bacteria like archaea. Bacillus and Clostridium produce o Eukaryotes: Include fungi, algae, endospores for survival under protozoa, animals, and plants. harsh conditions, which are highly resistant to environmental stresses. Viruses vs. Bacteria vs. Eukaryotes: 2. Cell Envelope Structures: Viruses: Acellular, obligate parasites that use host machinery. o Plasma Membrane: Prokaryotic Bacteria: Prokaryotic cells with unique plasma membranes are structures (e.g., peptidoglycan cell walls) phospholipid bilayers but lack that allow targeted antimicrobial treatment. sterols (except for Eukaryotes: More complex, with membrane- Mycoplasmataceae). bound organelles (e.g., fungi and parasites), making them more similar to human cells. o Cell Wall: The bacterial cell wall differs between Gram-positive and Key Points for Prelim Exam: Gram-negative bacteria: ▪ Gram-Positive: Thick Protozoa: Unicellular parasites, classified by peptidoglycan layer, with their locomotion. teichoic and lipoteichoic Fungi: Understand yeast vs. mold, and acids. dimorphism in fungi. Viruses: Acellular and dependent on host ▪ Gram-Negative: Thin cells for replication. peptidoglycan layer, surrounded by an outer  membrane containing o Plasma Membrane: Composed of a lipopolysaccharides (LPS). phospholipid bilayer with embedded proteins and o Acid-Fast Cell Wall: Found in cholesterol, which helps regulate organisms like Mycobacterium, it fluidity and stability. contains mycolic acid and is stained using acid-fast techniques. o Cell Wall: Present only in fungi, plants, and some protists; fungal 3. Surface Polymers: Capsules and slime layers cell walls contain chitin. provide protection and assist in adherence to surfaces. 3. Motility Organelles: o Flagella: Prokaryotic cells use o Cilia and Flagella: Used for flagella for movement, and the locomotion. Eukaryotic flagella and arrangement of flagella can help in cilia are structurally different from species identification. their prokaryotic counterparts and are typically longer (flagella) or o Pili/Fimbriae: Hair-like appendages numerous and shorter (cilia). used for adherence or exchange of genetic material during Summary of Key Differences: conjugation. Nucleus: Prokaryotes lack a true nucleus, Eukaryotic Cell Structure while eukaryotes have one. 1. Cytoplasmic Structures: Ribosome Size: Prokaryotes have 70S ribosomes, eukaryotes have 80S ribosomes. o Nucleus: Eukaryotes have a true nucleus with a nuclear membrane Cell Wall Composition: Prokaryotic cell walls containing DNA organized in differ based on Gram-positive/Gram- chromosomes. negative structure, while eukaryotes (like fungi) have specialized polysaccharide cell o Ribosomes: Eukaryotic ribosomes walls. are 80S in size (composed of 60S and 40S subunits), attached to the Organelles: Eukaryotes possess membrane- rough endoplasmic reticulum (ER). bound organelles, while prokaryotes do not. o Endoplasmic Reticulum (ER): The Bacterial Morphology rough ER is involved in protein synthesis, while the smooth ER Bacteria come in three basic shapes: synthesizes lipids. 1. Cocci (spherical): These can be found as o Mitochondria: The site of ATP individual cells, pairs (diplococci), chains production and contains its own (streptococci), or clusters (staphylococci). DNA. 2. Bacilli (rod-shaped): They vary in length and o Other Organelles: Lysosomes, may occur singly, in chains, or form peroxisomes, and chloroplasts (in palisades. Some bacilli are curved or have plant cells) serve specialized tapered ends. functions like degradation, 3. Spirochetes (spiral): These bacteria vary in protection, and photosynthesis. length and in the number of helical turns. 2. Cell Envelope Structures: Some species exhibit variability in size and shape (pleomorphism). Staining Techniques o A negative stain used to visualize capsules, particularly of Stains are used to color or highlight bacteria and help Cryptococcus neoformans, creating classify them based on their appearance under a a contrast between the capsule and microscope. the background. 1. Gram Stain: 8. Endospore Stain: o Divides bacteria into gram-positive o The Schaefer-Fulton method is (purple) and gram-negative (pink) used to stain bacterial spores, based on their cell wall structure. where spores appear green and o Gram-positive bacteria retain the bacterial cells red or pink. primary stain (crystal violet), while gram-negative bacteria are decolorized and take up the counterstain (safranin). 2. Acid-Fast Stains: o Used for bacteria with waxy cell walls (e.g., Mycobacterium species). o Stains include the Ziehl-Neelsen or Kinyoun method using carbol fuchsin and acid-alcohol as a decolorizer. Acid-fast bacteria Microbial Growth and Nutrition retain the red stain. Bacteria have three major nutritional needs for 3. Acridine Orange: growth: o A fluorescent dye that binds to nucleic acids, staining both gram- 1. Carbon Source: Essential for making cellular constituents, accounting for about 50% of a positive and gram-negative bacterium's dry weight. bacteria. 2. Nitrogen Source: Necessary for synthesizing 4. Calcofluor White: proteins and nucleic acids, representing 14% of the dry weight. o Binds to fungal cell wall chitin and 3. Energy Source: Required for ATP production fluoresces under UV light, used to to perform cellular functions. visualize fungal structures. In addition, smaller amounts of phosphorus, sulfur, 5. Methylene Blue: and various metal ions are crucial for enzymatic activity and other cellular processes. o Primarily stains Corynebacterium diphtheriae and is a counterstain Nutritional Requirements for Growth for acid-fast staining. 6. Lactophenol Cotton Blue: Bacteria are classified into two main groups based on their nutritional needs: o Stains fungal cell walls and is used for observing fungal morphology. Autotrophs (Lithotrophs): These bacteria can grow using carbon dioxide as their sole 7. India Ink: carbon source, requiring only water and  inorganic salts. They obtain energy either o Thermophiles (50° to 60° C) through photosynthesis (phototrophs) or by Most bacteria associated with oxidizing inorganic compounds humans are mesophiles, incubated (chemolithotrophs). at approximately 35° C. Heterotrophs: These bacteria need more 3. Gaseous Composition: Oxygen requirements complex organic substances for growth, vary among bacteria: often using organic carbon sources like o Obligate Aerobes: Require oxygen glucose for both carbon and energy. Most for growth. bacteria in the human body are o Facultative Anaerobes: Can grow heterotrophs, but their nutritional with or without oxygen. requirements vary widely. For instance, o Obligate Anaerobes: Cannot grow some can utilize a range of organic in the presence of oxygen. compounds, while others, like Haemophilus o Microaerophilic Bacteria: Require influenzae, are fastidious and require lower oxygen levels for growth. additional growth factors. Bacterial Growth Types of Growth Media Bacteria replicate through binary fission, with the Bacterial growth media can be categorized as follows: time taken for a single cell to divide called generation time, which can be as short as 20 minutes for fast Minimal Medium: Contains simple and growers like E. coli or up to 24 hours for slower completely defined ingredients; not species like Mycobacterium tuberculosis. commonly used in diagnostic labs. Nutrient Media: More complex media made Growth Curve from meat or soybean extracts (e.g., nutrient broth). The growth of bacterial cultures can be represented Enriched Media: Contains added growth by a growth curve showing four phases: factors, such as blood or vitamins (e.g., blood agar). 1. Lag Phase: Bacteria prepare to divide. Selective Media: Contains additives that 2. Log Phase: Bacterial numbers increase inhibit some bacteria but allow others to logarithmically. grow (e.g., MacConkey agar). 3. Stationary Phase: Nutrient depletion causes Differential Media: Enables visualization of stable bacterial numbers. metabolic differences (e.g., distinguishes 4. Death Phase: Nonviable cells outnumber lactose fermenters on MAC). viable ones. Transport Medium: Preserves the viability of microorganisms during transportation Determination of Cell Numbers without promoting growth. In diagnostic laboratories, bacterial cell numbers can Environmental Factors Influencing Growth be determined by: Three key environmental factors affect bacterial Direct Counting: Estimates total bacteria but growth: does not distinguish between live and dead cells. 1. pH: Most pathogenic bacteria thrive at a Direct Plate Count: Counts viable cells by neutral pH (7.0 to 7.5). growing dilutions on agar plates. 2. Temperature: Bacteria are classified based Density Measurement: Correlates turbidity on their optimal growth temperature: of broth cultures to viable cell counts o Psychrophiles (10° to 20° C) (CFU/mL), useful in antimicrobial o Mesophiles (20° to 40° C) susceptibility testing. Bacterial Biochemistry and Metabolism o Converts glucose-6-phosphate to pyruvate; produces one NADPH Metabolism and uses one ATP. Microbial metabolism encompasses Anaerobic Utilization of Pyruvic Acid (Fermentation) biochemical reactions for breaking down organic compounds and synthesizing new Various fermentation pathways yield distinct molecules. end products: Energy generation occurs during substrate o Alcoholic: Ethanol. breakdown; enzyme activity regulates o Homolactic: Lactic acid. metabolic processes. o Heterolactic: Lactic acid, CO₂, Bacterial identification utilizes metabolic alcohols. differences, analyzing substrate utilization, end products, and pH changes. o Propionic Acid: Propionic acid. Fermentation and Respiration o Mixed Acid: Multiple acids (lactic, acetic, etc.); positive methyl red Fermentation: reaction. o Anaerobic process with organic o Butanediol: Acetoin, 2,3- compounds as electron acceptors. butanediol; positive Voges- o Less efficient than respiration; Proskauer reaction. produces NAD essential for the o Butyric Acid: Butyric acid and other Krebs cycle. products. o End products include lactate, Aerobic Utilization of Pyruvate (Oxidation) butyrate, ethanol, and acetoin. The Krebs cycle oxidizes pyruvate, Aerobic Respiration: generating energy (ATP) and carbon o More efficient; molecular oxygen skeletons for biosynthesis. serves as the final electron Carbohydrate Utilization and Lactose Fermentation acceptor. Ability to ferment various sugars is critical o Anaerobic respiration uses for microbial identification. alternative electron acceptors (nitrate, sulfate). Lactose requires two enzymes for fermentation (transport and breakdown). Biochemical Pathways from Glucose to Pyruvic Acid Lactose fermenters can also ferment 1. Embden-Meyerhof-Parnas (EMP) Pathway: glucose. o Main pathway for glucose Bacterial Genetics Summary conversion to pyruvate; generates NADH and ATP; anaerobic. Historical Background 2. Pentose Phosphate Pathway: Key Discoveries: DNA was discovered by o Alternative pathway producing Frederick Miescher (1869), characterized by ribulose-5-phosphate and NADPH; Phoebus A. T. Levine (1920s), and its helical used by heterolactic fermenters. structure was revealed by Rosalind Franklin. Watson and Crick defined the three- 3. Entner-Doudoroff Pathway: dimensional structure in the 1950s. DNA and RNA Structure Transposons: Larger elements that carry additional genes, often drug-resistance DNA Structure: genes. o Double helix composed of Mutations deoxynucleotides: phosphate group, five-carbon sugar Changes in DNA code affecting protein (deoxyribose), and nitrogenous synthesis; can be silent, point mutations, or bases (A, T, C, G). frameshift mutations. o Bases pair: A with T and C with G, Occur spontaneously or due to external connected by hydrogen bonds. agents, with rates of 1 in 10⁹ and 1 in 10⁷ cells respectively. o Antiparallel strands run 3' to 5' and 5' to 3'. Genetic Recombination and Gene Transfer Mechanisms RNA Structure: 1. Transformation: Uptake of free DNA into a o Single-stranded, contains ribose, bacterial cell, integrating into the genome. and replaces thymine (T) with uracil (U). 2. Transduction: Gene transfer via bacteriophages; involves lytic and lysogenic Genetic Terminology cycles. Genotype: Genetic potential of DNA; 3. Conjugation: Direct transfer of DNA includes constitutive, inducible, and silent between bacteria through a pilus; involves genes. plasmids and chromosomal genes. Phenotype: Observable characteristics Restriction Enzymes produced by the genome. Cut foreign DNA at specific sites; bacteria Gene Flow: From DNA to mRNA to proteins; protect their own DNA through methylation. involves replication, transcription, and translation. Key Points to Remember Bacterial Genome Prokaryotes (bacteria) lack membrane- bound nuclei; eukaryotes do not. Consists of a single, circular, supercoiled dsDNA. Bacteria are classified by Gram stain (positive or negative). Contains genes that code for proteins and regulatory regions for transcription control. Bacterial spores form as a survival mechanism. Extrachromosomal DNA Elements Gram-negative bacteria possess LPS, which Plasmids: Circular dsDNA that carry genes can cause fever and shock. for traits like antimicrobial resistance; can replicate independently and be transferred Bacteria generate energy through via conjugation. fermentation and respiration. Mobile Genetic Elements Origin of Microbial Biota Insertion Sequences (IS): Small pieces of Colonization vs. Transient Organisms DNA that can disrupt genes.  Colonization: Microorganisms that o Resident Microbiota: successfully establish themselves in a Microorganisms that colonize an specific area of the host are referred to as area for extended periods. colonizers. They become predominant o Transient Microbiota: organisms in that niche. Microorganisms that temporarily inhabit a site, eliminated by Transient Organisms: Some microorganisms immune defenses or competition are transient and do not establish a lasting with resident biota. presence; they are eliminated either Acute Carriers: Transient carriers, such as through the host's immune defenses or those who harbor Neisseria meningitidis for a short period during outbreaks. competition with resident microbiota. Chronic Carriers: Long-term carriers, like individuals with Salmonella Typhi, who may Host-Microbe Relationships harbor the pathogen for years without symptoms. Once established, microorganisms can develop various relationships with their host, which can be categorized as: Factors That Determine the Composition of the Usual Microbial Biota 1. Symbiosis: A general term for two organisms living together. Within this category, there The composition of microbial biota at specific body are: sites is influenced by several factors: o Mutualism: Both organisms’ benefit. For example, Lactobacilli in 1. Nutritional Factors: The availability and type the urogenital tract of women of nutrients affect the microbial provide protection against colonization. For instance, areas with more pathogens while deriving nutrients moisture host greater microbial diversity. from the host. 2. Environmental Factors: Conditions such as o Commensalism: One organism skin lipids, bile, and pH determine which benefit while the other is microorganisms can thrive. For example: unaffected. Proteus mirabilis is an o The female genital tract has a pH of example of a commensal in the 4.0 to 5.0, which limits the survival gastrointestinal tract, where it of many bacteria. exists without causing harm. o The intestinal biota differs o Parasitism: One organism benefit at significantly between breast-fed the expense of the host. An and formula-fed infants, with example is Entamoeba histolytica, breast milk promoting higher levels which causes intestinal ulcers and of Bifidobacterium due to its dysentery by deriving nutrients lactose content. from the host. 3. Oxidation-Reduction Potential: Anaerobic conditions favor fermentation organisms, Characteristics of Indigenous Microbial Biota such as those found in gingival crevices. Indigenous microbiota refers to microorganisms that Changes Influencing Microbial Biota are regularly found on or within healthy individuals. The characteristics of these biota include: The microbial biota can change due to various factors, including: Diversity: Different body sites host different microbial communities, influenced by local Age: Immune response effectiveness conditions such as moisture, pH, and typically declines with age, increasing available nutrients. susceptibility to opportunistic infections. Resident vs. Transient Microbiota:  Nutritional Status: Changes in diet can alter Upper Respiratory Tract: Includes the the microbial community. mouth, nasopharynx, oropharynx, and Disease States: Certain health conditions can larynx. shift microbial populations. Lower Respiratory Tract: Comprises the Antimicrobial Therapy: Antibiotics can trachea, bronchi, and lungs, which are disrupt established microbiota, leading to an typically sterile due to protective increase in opportunistic infections. mechanisms like ciliary action and mucus movement. Normal Microbiota of the Skin Colonizing Organisms: o Upper Respiratory Tract: The skin serves as a barrier against infection through Predominantly colonized by several protective mechanisms, including: viridans streptococci (e.g., Streptococcus mitis, S. mutans, S. anginosus, S. sanguinis), along with Physical Barriers: Preventing direct contact Moraxella catarrhalis, Neisseria between pathogens and underlying tissues. spp., and diphtheroids. Anaerobes Chemical Defenses: Fatty acids and thrive in gingival crevices. lysozymes inhibit microbial growth. o Oropharynx: Hosts a diverse Desquamation: Shedding of dead skin cells mixture of streptococci, including helps eliminate bacteria. viridans group streptococci and Distribution: Microbial populations are opportunistic pathogens like S. concentrated in moist areas (e.g., armpits, aureus. groin). Common Genera: Staphylococcus Normal Microbiota of the Gastrointestinal Tract epidermidis and Propionibacterium spp. are prevalent, particularly in sebaceous glands and hair follicles. Propionibacterium acnes The gastrointestinal tract includes the esophagus, can be found deeper in the sebaceous stomach, small intestine, and colon, and it is glands, often contaminating clinical equipped with various defense mechanisms. specimens. Diversity: Despite strong defenses, over Normal Microbiota of the Oral Cavity 35,000 bacterial species may inhabit the GI tract. Microbial Population Distribution: The oral cavity harbors a rich microbial environment, o Esophagus: Contains very few primarily characterized by the presence of bacteria. microorganisms (about 10 microbes per gram). Dominant Genus: Streptococcus, with high o Stomach: Gastric juices (pH 2) densities leading to significant plaque eliminate most bacteria, but some formation (up to 1011 streptococci per gram (e.g., Helicobacter pylori) can of plaque). survive by adhering to the stomach Anaerobic Conditions: Plaque creates a low lining. oxidation-reduction potential, facilitating the o Small Intestine: Fewer growth of strict anaerobes in crevices and microorganisms than the colon, interstitial spaces between teeth. with some organisms moving here after surviving gastric acid. Normal Microbiota of the Respiratory Tract o Colon: Contains approximately 1012 bacteria per gram and is home to The respiratory tract is divided into the upper and over 70% of the body's total lower sections: microbiota, primarily obligate anaerobes such as Bacteroides, Clostridium, and Prevotella. Effects of Antibiotics: Antibiotic treatment can alter and activating cell-mediated the gut microbiota, leading to overgrowth of immunity. opportunistic pathogens like Clostridium difficile and Candida albicans. Pathogenesis and Virulence Normal Microbiota of the Genitourinary Tract 1. Pathogenicity: o Refers to a microbe's ability to The genitourinary tract generally remains sterile in cause disease, which can vary internal structures, although some organisms can between true pathogens (e.g., colonize the urethra and vagina. Yersinia pestis, Bacillus anthracis) and opportunistic pathogens. Urethra: Colonized by skin organisms, 2. Iatrogenic Infections: particularly in women. o These are infections that occur due Vaginal Microbiota: Influenced by hormonal to medical interventions (e.g., changes, particularly estrogen. In catheter insertion), highlighting the childbearing years, lactobacilli dominate, need for strict aseptic techniques. helping to maintain a low pH and inhibiting pathogen growth. In prepubescent and Routes of Transmission postmenopausal women, a different microbiota composition prevails. 1. Airborne Transmission: o Infectious agents can spread Role of Microbial Biota in Infectious Disease through respiratory secretions, with small particles remaining 1. Symbiotic Relationships: airborne for extended periods. o Microbial biota usually benefits the o Common infections include host by aiding in digestion, streptococcal throat and synthesizing vitamins, and pneumonia. preventing pathogen colonization. 2. Food and Water Transmission: o Some members can become o Gastrointestinal infections often opportunistic pathogens if the arise from ingesting contaminated host’s immune system is food or water. compromised or if their normal o Pathogens can either produce habitat is altered (e.g., after trauma toxins before ingestion or cause or surgery). tissue damage after infection (e.g., 2. Opportunistic Infections: E. coli, Vibrio cholerae). o Organisms like H. influenzae and S. 3. Close Contact: epidermidis are typically harmless o Many infections spread via direct but can cause serious infections in salivary contact (e.g., kissing) or immunocompromised individuals. skin contact (e.g., herpes, HPV). o Changes in the microbial 4. Bite Wounds: community (due to factors like o Animal and human bites can antibiotic use) can lead to introduce oral biota into wounds, infections by previously benign leading to serious infections. organisms (e.g., C. albicans). 5. Arthropod Vectors: 3. Immune System Development: o Many infectious diseases (e.g., o Exposure to microbes is crucial for malaria, Lyme disease) are developing a competent immune transmitted through bites from system; germ-free animals show infected arthropods. poor immune function. 6. Zoonoses: o The microbial biota plays a role in o These diseases are transmitted stimulating antibody production from animals to humans through various means (bites, contact with  secretions, etc.), emphasizing the Receptors: Host cells must have receptors interconnection between human that match the adhesins for effective and animal health. binding. Virulence Intracellular Survival Definition: The relative ability of a Intracellular Pathogens: Some organisms can microorganism to cause disease, often survive and replicate within phagocytic cells, measured by the infective dose needed to avoiding destruction (e.g., Chlamydia, establish infection. Mycobacterium). High vs. Low Virulence: Organisms requiring Antigenic Variation: Some pathogens shift a low infective dose (e.g., Shigella spp. with their surface antigens to evade host 100 organisms) are considered more antibodies. virulent than those needing a higher dose. Toxin Production Microbial Virulence Factors Exotoxins: These are potent toxins secreted Mechanisms of Infection: Microorganisms by bacteria, composed of two subunits (one have evolved various virulence factors that binding, one toxic). enable them to persist in a host and evade Endotoxins: Part of the outer membrane of immune defenses. These include: gram-negative bacteria, responsible for o Capsules: Many bacteria have systemic effects like fever and shock when polysaccharide capsules that inhibit released in large amounts. phagocytosis. o Toxins: Substances that damage Effects of Endotoxins host tissues, with exotoxins being secreted and endotoxins being part Systemic Response: Endotoxins trigger the of the gram-negative bacterial cell release of cytokines, leading to severe wall. inflammatory responses, including fever and hypotension, and can result in septic shock. Resistance to Phagocytosis Summary of Exotoxins vs. Endotoxins Phagocytes: Cells like macrophages and neutrophils ingest and destroy pathogens. Exotoxins: Evasion Techniques: o Secreted by both gram-negative o Capsules: Masking cell surface and gram-positive bacteria. structures recognized by o Specific action on host cells. phagocytes. o Can be inactivated by heat. o Protein A: In S. aureus, this protein Endotoxins: interferes with antibody binding, o Only found in gram-negative preventing opsonization. bacteria. o Toxin Production: Some bacteria o Non-specific and less potent. release substances (e.g., o Heat-stable and released upon cell leukocidins) that kill phagocytes. lysis. Adhesion to Host Cells Host’s Resistance Factors against Infection Adhesins: Surface structures (fimbriae/pili, 1. Physical Barriers polysaccharides) that enable pathogens to attach to host cells, a critical step for Skin: The primary mechanical barrier to infection. infection, with its stratified and cornified  epithelium preventing most microorganisms Some produce substances like bacteriocins from penetrating. to inhibit related bacteria, contributing to Mucous Membranes: Generally, allow entry host health by synthesizing essential for pathogens but are not easily penetrated nutrients. by intact skin. 5. Phagocytosis 2. Cleansing Mechanisms Key Process: Essential for the host's defense Desquamation: Continuous shedding of the against infections; involves the ingestion and outer layer of skin helps remove destruction of pathogens by phagocytes microorganisms. (PMNs, macrophages, monocytes). Fluids: Various body fluids (tears, mucus) Chemotaxis: Directed migration of have antimicrobial properties and physically phagocytes to infection sites, stimulated by cleanse surfaces by washing away various substances. pathogens. Attachment and Ingestion: Phagocytes use o Eyes: Tears contain IgA and antibodies and complement proteins for lysozyme to protect against opsonization to facilitate attachment and infections. ingestion of pathogens. o Respiratory Tract: Mucus traps Killing Mechanism: Following ingestion, particles and microorganisms; cilia phagocytes undergo a metabolic burst that sweep them toward the throat for enhances their ability to kill pathogens expulsion or swallowing. through the production of reactive oxygen o Gastrointestinal Tract: Stomach species and various enzymes. acid kills most bacteria; mucous secretions and peristalsis prevent 6. Inflammation attachment of organisms. o Genitourinary Tract: Urine flow A nonspecific response to injury or infection helps cleanse and maintain low characterized by increased blood flow, microbial populations. swelling, and accumulation of phagocytes. Mediators: Chemicals released during 3. Antimicrobial Substances inflammation lead to tissue repair and removal of pathogens. Lysozyme: An enzyme that breaks down bacterial cell walls and is found in various 7. Immune Responses bodily fluids. Secretory IgA: Antibodies in mucous The immune system encompasses both secretions that enhance phagocytosis and innate (natural) and adaptive (specific) neutralize pathogens. responses to pathogens. β-Lysins: Proteins released during The balance of immune responses is coagulation that are effective against gram- complex and influenced by various factors, positive bacteria. including the host's health and genetic Interferon: A protein that inhibits viral makeup. replication and enhances immune responses. Innate Immunity 4. Indigenous Microbial Biota Definition: The first line of defense against pathogens; non-specific and does not Nonpathogenic microorganisms compete require prior exposure. with pathogens, reducing their chance of Components: colonization.  1. Physical and Chemical Barriers: but release lymphokines that affect Skin, mucous membranes, other immune cells. secretions (like saliva and tears). o Effective against intracellular 2. Blood Proteins: Act as mediators in pathogens, with a focus on infection. destroying infected host cells. 3. Cellular Mechanisms: Includes phagocytic cells (neutrophils, Pathogen-Specific Responses macrophages) and natural killer cells. Extracellular Bacteria: Primarily managed by Function: Rapid response to pathogens, antibodies (humoral immunity) and primarily effective against extracellular phagocytosis. bacteria; plays a minor role against Intracellular Pathogens: Managed intracellular pathogens and viruses. predominantly through T cell activity and Recognition: Utilizes pattern recognition lymphokines. receptors (PRRs), such as toll-like receptors Viruses: Both humoral and cell-mediated (TLRs), to identify microbial components. responses are important; neutralizing antibodies can prevent infections. Adaptive Immunity Fungal and Parasitic Infections: Primarily controlled by cell-mediated immunity, with Definition: A specific immune response that little protective role from antibodies. develops memory and is tailored to distinct antigens. Mechanisms by Which Microbes May Overcome Host Key Players: Lymphocytes (B cells and T Defenses cells). B Cells: Produce antibodies in response to 1. Tolerance and Immunosuppression: antigens; responsible for humoral immunity. o Some pathogens induce tolerance, T Cells: Mediate cellular immunity and can where the host fails to mount an target infected host cells directly. adequate immune response to Immunologic Memory: The ability to certain antigens, often because respond more vigorously to previously they are perceived as “feeble encountered antigens. antigens.” This can occur if the host is exposed during fetal Immune Response Mechanism development or in infancy, as seen with the rubella virus, which can Humoral Immunity: persist without causing overt o B cells differentiate into plasma disease. cells that secrete antibodies o Additionally, some microbes (immunoglobulins). actively suppress the host’s o Antibody types (IgG, IgM, IgA, IgD, immune response. For instance, IgE) vary in structure and function. viruses such as Epstein-Barr virus o Primary vs. Secondary Responses: and cytomegalovirus can reduce T- ▪ Primary Response: Initial cell or antibody responses to other production of IgM antigens. The most notable followed by IgG. example is HIV, which targets and ▪ Secondary Response: destroys CD4+ T cells, Faster and stronger compromising the host’s defense increase in IgG levels upon against various pathogens. re-exposure. 2. Antigenic Variation: Cell-Mediated Immunity: o Certain microbes, like Borrelia o Primarily involves T lymphocytes, recurrentis, can change their which do not produce antibodies surface antigens during an infection, allowing them to evade  detection by the immune system. True pathogens are capable of causing This results in recurrent symptoms, disease in any individual. as the immune system responds to The host's defense mechanisms consist of the initial antigen but fails to physical barriers, components of the innate recognize the modified version in immune system (like phagocytes and subsequent waves of infection. cytokines), and the adaptive immune 3. Intracellular Survival: response. o Some pathogens, such as Brucella, Microbes possess various evasion strategies, Listeria, and mycobacteria, evade including avoiding phagocytosis, producing the immune response by surviving toxins, inducing tolerance, or varying surface within host cells, particularly antigens to escape recognition by the macrophages. By residing adaptive immune system. intracellularly, they become less accessible to antibodies and other Disinfection and Sterilization Overview immune defenses, thus promoting their survival and replication. Disinfection refers to processes that 4. Low-Avidity Antibodies: eliminate a defined range of o Even when the host produces microorganisms, including some spores, but antibodies, these may have low do not achieve complete sterility. avidity or weak antimicrobial Sterilization is the destruction of all forms of effects, which can impair the host's microbial life, including bacterial spores, and ability to control the infection is an all-or-nothing process. effectively. In such cases, the antibodies produced do not provide sufficient protection. Key Definitions 5. Evasion of Interferons: o Interferons play a critical role in the Antiseptics: Substances applied to skin to immune response by activating reduce bacteria, but they do not kill spores. immune cells and exhibiting Chemical and Physical Methods: Various antiviral properties. However, some methods exist for disinfection and viruses can evade these effects. For sterilization, which can be categorized based example, the vaccinia virus can on their application. inactivate interferon gamma, while other viruses may fail to induce Factors Influencing Killing Efficacy interferon production, allowing them to establish persistent 1. Types of Organisms: infections. o Microorganisms vary in resistance to disinfectants. For example, Key Points to Remember: bacterial endospores are particularly resilient due to their Humans are constantly exposed to protective structure, while lipid-rich microorganisms, starting from birth, and our viruses are more susceptible to body hosts a typical microbial biota detergents. influenced by environmental factors. 2. Number of Organisms (Bioburden): Some of these microbes can become o The total microbial load affects the opportunistic pathogens, causing disease effectiveness of disinfection; higher when the host’s immune system is numbers generally require longer compromised. exposure to disinfectants. The normal microbial biota also provides 3. Concentration of Disinfectant: protective benefits, such as competing with o The effectiveness of a disinfectant pathogens and priming the immune system. is concentration-dependent. Following manufacturer guidelines  is essential to ensure adequate exposure to infectious agents, including killing power. proper use and disposal of disinfectants. 4. Presence of Organic Material: o Organic matter can inactivate Regulatory Considerations disinfectants and shield microorganisms, necessitating The use of disinfectants and antiseptics is thorough cleaning before subject to regulatory processes to ensure disinfection. safety and efficacy in health care settings. 5. Nature of Surface: o The material of the surface to be Overview of Disinfection and Sterilization disinfected can affect the choice of disinfectant, as some methods may damage certain surfaces. Sterilization vs. Disinfection: 6. Contact Time: o Sterilization is the complete o The time a disinfectant or sterilant destruction of all forms of life, remains in contact with a surface is including spores, through physical crucial. Insufficient contact time or chemical methods. can lead to ineffective disinfection. o Disinfection reduces the number of 7. Temperature: viable microorganisms to a level o Disinfectant efficacy often considered safe, but does not kill all increases with temperature; spores. however, extreme temperatures o Antiseptics are used on living can reduce effectiveness. tissues to reduce the number of 8. pH: bacteria but do not kill spores. o The pH of the disinfecting solution and the material being disinfected Factors Influencing Disinfection and Sterilization can influence the efficacy of the disinfectant. 1. Types of Organisms: Resistance varies; 9. Biofilms: prions are the most resistant, while o Biofilms provide a protective enveloped viruses are more susceptible. environment for microorganisms, 2. Number of Organisms: Higher microbial load making them more resistant to requires longer exposure to disinfectants. disinfectants. This often requires 3. Concentration of Disinfectant: Proper adjustments to disinfectant concentration is crucial for effective concentration and contact time. microbial killing. 10. Compatibility of Disinfectants: 4. Presence of Organic Material: Organic o The interaction between multiple matter can inactivate disinfectants, so disinfectants can affect their surfaces must be cleaned beforehand. efficacy. Some combinations can 5. Nature of the Surface: Material compatibility inactivate one another, such as affects choice of disinfection methods. bleach and quaternary ammonium 6. Contact Time: Longer contact times increase compounds. effectiveness. 7. Temperature and pH: Optimal conditions Practical Applications enhance the activity of disinfectants. 8. Biofilms: These can protect microorganisms, Common Disinfectants: Familiarity with requiring more rigorous disinfection common disinfectants and their appropriate methods. use is essential for effective microbial 9. Compatibility of Disinfectants: Some control in health care settings. disinfectants can inactivate each other. Laboratory Safety Guidelines: Laboratories must develop protocols to minimize Historical Contributions  Ignaz Semmelweis: Advocated for Mechanism: Kills microorganisms through handwashing to reduce maternal mortality oxidative effects of hypochlorous acid in rates by preventing the transfer of water. pathogens from cadavers to patients. Uses: Effective as surface disinfectants and Joseph Lister: Introduced antiseptic for water disinfection. techniques in surgery, significantly reducing Limitations: Not sporicidal, inactivated by infection rates. organic matter, corrosive, and requires long exposure time. Common Disinfectants and Antiseptics Quaternary Ammonium Compounds Alcohols: Effective against most bacteria and some viruses but not spores; used primarily Characteristics: Cationic surfactants that as antiseptics. reduce surface tension in liquids. Aldehydes: Mechanism: Disrupt cell membranes, o Formaldehyde: Used for causing leakage of cell contents. disinfection but has irritant and Limitations: Effectiveness reduced by hard carcinogenic properties. water, soaps, and organic matter; not o Glutaraldehyde: A high-level effective against certain bacteria (e.g., disinfectant that remains active in Pseudomonas aeruginosa). organic matter; effective against a broad spectrum of pathogens. Phenolics Halogens: o Iodophors: Used as antiseptics and Composition: Modified phenol molecules disinfectants; effective when with halogens or alkyl groups to enhance properly diluted. Examples include efficacy. povidone-iodine. Mechanism: Disrupt cell walls and precipitate proteins. Regulatory and Safety Considerations Uses: Commonly used in hospitals and household disinfectants. Disinfectants and antiseptics undergo a Limitations: Not sporicidal; effectiveness can regulatory process to ensure safety and be influenced by organic material. efficacy. Guidelines emphasize the importance of Chlorhexidine Gluconate (CHG) using the right concentrations and following manufacturer instructions to prevent Use: Approved for topical antiseptic use; infections effectively. binds strongly to skin. Mechanism: Disrupts microbial cell Application in Clinical Settings membranes. Spectrum: More effective against gram- Effective disinfection and antisepsis are positive bacteria; inactive against spores and critical in controlling nosocomial infections, some viruses. especially in healthcare environments where Advantages: Low toxicity, persistent activity the risk of contamination is high. for at least 6 hours. Chlorine and Chlorine Compounds Hexachlorophene Forms: Commonly used as hypochlorite Mechanism: Disrupts bacterial electron (e.g., sodium hypochlorite, calcium transport and membranes. hypochlorite). Limitations: Primarily effective against gram- positive bacteria; associated with severe toxicity. Chloroxylenol (PCMX) Label Claims: Must specify effectiveness against certain microorganisms; general Mechanism: Disrupts cell walls and disinfectant claims must be backed by inactivates enzymes. broad-spectrum efficacy. Limitations: Intermediate-acting with low persistent effect; less effective against gram- FDA Regulations on Chemical Skin Antiseptics negative bacteria. Approval Processes: Manufacturers can Triclosan apply through NDA or OTC drug review. Categories: Products classified based on Mechanism: Disrupts cell walls; shows good safety and effectiveness; specific efficacy activity against a variety of pathogens. testing required. Concerns: Associated with potential thyroid hormone disruption and antibiotic Hand Hygiene resistance; regulatory scrutiny over long- term safety. Hygienic Handwashing: Aimed at removing transient biota; essential for preventing Heavy Metals infection. Surgical Hand Scrubs: Designed to reduce Examples: Silver nitrate used for preventing both transient and resident biota; FDA gonococcal conjunctivitis. requires significant bacterial reduction Limitations: Rarely used due to toxicity; slow within a short time. bactericidal action; largely replaced by safer Preoperative Skin Preparation: Must alternatives. effectively reduce microorganisms on surgical sites; also aims for persistence of Gaseous Disinfectants antimicrobial activity. Ethylene Oxide: Used for sterilization; Historical Context effective against spores and vegetative cells; requires careful control of concentration, Pre-1980s Practices: Laboratory safety temperature, and humidity. practices were lax; common behaviors Hydrogen Peroxide: Active against all types included mouth pipetting and eating or of microorganisms; used in vapor form for drinking in the lab, despite being sterilization. discouraged. Peracetic Acid: Effective sterilant in gaseous Impact of AIDS: The arrival of AIDS, which form; similar action to hydrogen peroxide. had a near 100% mortality rate, prompted a significant re-evaluation of safety protocols Combination of Hydrogen Peroxide and Peracetic and employee risk for laboratory-acquired Acid infections (LAIs). This shift marked a move away from the mentality of "What you don't know can't hurt you." Advantage: Shorter contact time required for effective sterilization compared to individual components. Shift in Safety Culture EPA Regulations on Chemical Surface Disinfectants Increased Awareness: The awareness of biological hazards led to a prioritization of safety for laboratory personnel. Registration: Regulated under the Federal Reevaluation of Practices: Continuous Insecticide, Fungicide, and Rodenticide Act; reassessment of work practices is necessary requires laboratory and toxicity data. to enhance safety measures. Risk Factors provided, and a designated safety officer should oversee safety practices. Increased Infection Risk: Studies indicate Commitment to Safety: A safe work that laboratory workers are at a higher risk environment requires the active of infections compared to the general participation and commitment of all population. personnel. Untraceable Exposures: Laboratory exposures often occur without a clear Safety Program Components connection to specific events, making it crucial to maintain rigorous safety protocols. A comprehensive safety program should include the following elements: CDC Guidelines Biological Hazards: Conduct biological risk 2012 Guidelines: The CDC published assessments and develop safety procedures “Guidelines for Safe Work Practices in for handling biological materials. Human and Animal Medical Diagnostic Chemical and Radioactive Safety: Provide Laboratories” to promote safer work guidelines for the safe handling, storage, environments. and disposal of chemicals and radioactive o Goals: The guidelines aim to substances. improve safety practices, Emergency Procedures: Outline protocols encourage consideration of safety for emergencies like fires, natural disasters, issues, and foster a culture of and bomb threats. safety among laboratory staff. Initial and Ongoing Training: Conduct initial safety training for all employees, with Scope of Safety Measures annual updates. Manual Handling Training: Teach safe lifting Comprehensive Protection: Safety in clinical and moving techniques for heavy objects laboratories encompasses various types of and patients. hazard protection: o Biological Hazards: Risks from OSHA Regulations infectious agents. o Chemical Hazards: Risks from Hazard Awareness: Laboratory personnel hazardous substances. must recognize they work in a hazardous o Electrical Hazards: Risks associated environment, with potential biological, with electrical equipment. chemical, radiologic, or physical hazards. o Radioactive Hazards: Risks from OSHA Mission: The Occupational Safety and radioactive materials. Health Administration (OSHA) aims to o Fire Hazards: Risks related to protect workers and has regulations flammable materials and fire applicable to clinical laboratories. safety. Bloodborne Pathogens Standard: Established in 1991 and revised in 2001, this General Laboratory Safety standard outline employer responsibilities to protect employees from bloodborne Responsibility: Safety in clinical laboratories pathogens. is a shared responsibility among institutions, laboratory directors, managers, and Exposure Control Plan employees. Training: Employees must receive annual Employers must develop an exposure control plan training on current safety regulations and that includes: procedures. Safety manuals should be  Task Determination: Identify tasks that pose Closed Tube Sampling: To minimize occupational hazards. exposure. Incident Investigation: Implement plans to Safety Devices: Use of safety needles, investigate exposure incidents and prevent eyewash stations, and barriers. recurrence. Laboratory Environment: Maintenance of Compliance Methods: Establish compliance negative air pressure and limited access to methods for standard precautions, including laboratories. engineering and work practice controls, personal protective equipment (PPE), and Work Practice Controls waste disposal guidelines. Training Programs: Ensure comprehensive These controls modify how tasks are performed to training for all employees. reduce exposure risks: Standard Precautions Prohibitions: No mouth pipetting, eating, or drinking in laboratories. Updated in 1996 from universal precautions, Cleaning Protocols: Disinfection of standard precautions are essential for preventing workstations and safe specimen transport. bloodborne infections and include: Handwashing: Frequent and thorough hand hygiene practices. Handwashing: Essential after handling blood, body fluids, and contaminated items. Personal Protective Equipment (PPE) Use of Gloves: Required when handling any potentially infectious materials. Provision and Maintenance: Employers must Protective Gear: Masks, eye protection, and provide PPE such as gloves, masks, and lab laboratory coats must be worn when there's coats. a risk of exposure. Proper Usage: PPE must fit well, be used Sharps Disposal: Implement safe disposal whenever there’s a risk of exposure, and be methods for needles and other sharps in removed before leaving work areas. puncture-resistant containers. Fit Testing: Respirators should be fit-tested Environmental Control: Regular cleaning and for adequate protection against airborne disinfection of surfaces. hazards. Transmission-Based Precautions Biological Risk Assessment Overview These precautions are added for patients known or Conditions for Infection: An infection, suspected to be infected with infectious agents and including laboratory-acquired infections include: (LAIs), requires: 1. A susceptible host. Contact Precautions: For agents spread by 2. An infectious agent with a direct or indirect contact. transmission route to the host. Droplet Precautions: For agents spread 3. Sufficient concentration of the through respiratory secretions. agent to cause disease. Airborne Precautions: For agents that remain infectious in the air. Sources of Biological Hazards Engineering Controls Patient Specimens: Risks arise from processing patient specimens and handling Defined by OSHA, engineering controls aim to isolate growing cultures of microorganisms. or remove hazards from the workplace, such as: Routes of LAIs: Common routes include: o Parenteral: Needlesticks or contaminated sharps.  o Skin/Mucous Membranes: Spills or 5. Proper PPE Usage: Always wear laboratory splashes. coats and appropriate PPE; do not leave the o Ingestion: Mouth pipetting or lab wearing PPE. handling contaminated items. 6. Hand Hygiene: Ensure handwashing facilities o Inhalation: Infectious aerosols. are available, and wash hands before leaving the laboratory. High-Risk Infectious Agents Biological Risk Assessment Process Tuberculosis (M. tuberculosis): Risk of exposure through aerosols in sputum Biological risk assessment is critical in identifying processing. hazardous characteristics of infectious agents and Brucella spp. and Francisella tularensis: developing safety measures: Transmitted via aerosol during specimen handling. Assess Hazard Characteristics: Consider the Coccidioides immitis: Infectious fungus that agent's ability to infect and cause disease, can spread in an open lab environment. virulence, availability of treatment, and Bloodborne Pathogens: preventive measures (like vaccines). o Hepatitis B Virus (HBV): High WHO Risk Group Classification: transmission risk through o Risk Group 1: Low risk, unlikely to needlestick injuries; historically cause disease. caused thousands of infections o Risk Group 2: Moderate risk, among healthcare workers. treatable, limited spread. o HIV and Hepatitis C Virus: Also pose o Risk Group 3: High individual risk, risks through contaminated treatable, low community spread. specimens. o Risk Group 4: High risk, readily transmissible, limited treatment. Importance of Awareness Risk Assessment Steps (CDC Guidelines) Laboratory personnel must be aware of the risks associated with various infectious agents and the 1. Identify Hazards: Determine hazards potential for LAIs. Proper safety practices and associated with the infectious agent. protocols are essential to minimize exposure. 2. Identify Exposure Activities: Recognize activities that might lead to exposure. Recommendations to Enhance Safety 3. Consider Personnel Experience: Evaluate competencies and experience of laboratory Following an outbreak of Salmonella typhimurium staff. infections linked to laboratories, several 4. Evaluate and Prioritize Risks: Assess recommendations were made: likelihood and severity of potential LAIs. 5. Develop and Implement Controls: Create 1. Awareness of Hazards: Personnel must and evaluate controls to minimize exposure