Microbiology Chapter 4

What two things does taxonomy enable scientists to do?

1. organize large amounts of information about organisms 2. make predictions based on knowledge of similar organisms

Five types of information that microbiologists commonly use to distinguish among microorganisms

1. physical characteristics 2. biochemical tests 3. serological tests 4. phage typing 5. analysis of nucleic acids

Two types of probe microscopes

1. scanning tunneling microscopes 2. atomic force microscopes

Who compared nucleotide sequences of rRNA subunits?

Carl Woese

Why must a smear be fixed to a slide?

Fixation causes the specimen to adhere to the glass so that it does not easily wash off during staining

Name the two types of special staining

1. negative (capsule) stain 2. flagellar stain

Type of differential stain that differentiates Gram-positive and Gram-negative bacteria, which is typically the first step in their identification smear soak in violet dye rinse flood smear with iodine solution rinse rinse with ethanol and acetone rinse flood smear with safranin rinse blot dry

Gram stain procedure

After Linnaeus, later taxonomic approach based organisms into five kingdoms. What are they?

1. Animalia 2. plantae 3. fungi 4. Protista 5. prokaryotae the goal is to understand the relationship amoung groups of organisms

Usually able to identify protozoa, fungi, algae, and parasitic worms solely on their morphology can also use physical appearance of the bacterial colony, size and shape of the individual bacterial cells, or the presence of endospores or flagella.

Physical characteristics

Hierarchy Taxon Name the eight phyla in order from most broad to most specific

1. Domain 2. Kingdom 3. Phylum 4. Class 5. Order 6. Family 7. Genus 8. Species

What three domains are determined by ribosomal nucleotide sequences?

1. Eukarya 2. bacteria 3. archaea

Two popular stains for histological specimens

1. Gomori methenamine silver (GMS) 2. Hematoxylin and eosin (HE)

Name the four most common types of differential stains

1. Gram stain 2. acid-fast stain 3. endospore stain 4. histological stain

What are the four general principles of microscopy?

1. Wavelength of radiation 2. magnification 3. resolution 4. contrast

Two types of biochemical tests for identifying bacteria

1. carbohydrate utilization test 2. hydrogen sulfide test

Type of probe microscope that measures the deflection of a laser beam aimed at the tip of a probe that travels across the surface of the specimen.

atomic force microscope

Two types of bight-field microscopes

1. simple 2. compound

Two types of electron microscopes

1. transmission electron microscopes 2. scanning electron microscopes

Light microscopes cannot resolve structures closer than _____ nm.

200

Cells in the three domains differ by other ______.

characteristics

Linnaeus classified organisms based on _____ in common.

characteristics

Transmission electron microscopy uses _____ containing heavy metals that absorb electrons.

chemicals

Microbiological stains contain _____.

chromophore

The assigning of organisms to taxa based on similarities used in taxonomy

classification

Taxonomy consists of _____, _____, and _____.

classification, nomenclature, identification

The emphasis on comparison of organisms' genetic material led to the proposal to add _____ to the classification of organisms.

domain

Microscope that use electrons traveling as waves with shorter wavelengths than visible light, so the resolving power is greater can view ultrastructural details require specimens to be in a vacuum so cannot be used to examine living microbes

electron microscope

Typical magnification 1000X to 100,000X; resolution to .001nm.

electron microscope

The practical science of determining that an isolated individual or population belongs to a particular taxon used in taxonomy

identification

One thousandth of a micrometer (um) is a _____.

nanometer (nm)

Visible light is made of _____ bands of wavelengths of radiation.

narrow

Type of special stain where background is dark and reveals bacterial capsules

negative (capsule) stain

The naming of organisms (and the rules involved) used in taxonomy each species has a genus name and a specific epithet

nomenclature

Why isn't the Gram stain utilized to stain Mycobacterium?

Cell walls of mycobacterium are composed of waxy materials that repel the water-based dyes of the Gram stain

Series of lenses for magnification light passes through specimen into objective lens have one or two ocular lenses most have condenser lens which direct light through the specimen. uses oil immesion

Compound microscope

Type of microscope that uses fluorescent dyes; uses UV lasers to illuminate fluorescent chemicals in a single plane; resolution is increased because light passes through pinhole aperture; computer constructs 3-D image from digital images.

Confocal microscope

_____ is important in determining resolution.

Contrast

Best for observing pale objects only light rays scattered by specimen enter objective lens specimens appear light against a dark background increases contrast and enables observation of more details uses a dark-field stop in the condenser that prevents light from directly entering the objective lens

Dark-field microscopes

Acidic dyes stain _____ structures.

alkaline (basic) by ionically bonding to them it works best in acidic environments

Total magnification =

objective lens x ocular lens

Procedure involves spreading a solution across a solid surface of growth medium then adds small drops of different phages that results in if the phage infects and kills the bacteria a clear plaque will form

phage typing

Viruses that infect and usually destroy bacterial cells; very specific to which host they can infect

phages

Microscope that examines living organisms or specimens that would be damaged/altered by attaching them to slides or staining contrast is created because light waves are out of phase

phase microscopes

What are the two types of phase microscopes?

phase-contrast differential interference contrast

Type of phase microscope that is the simplest produces sharply defined images particularly useful for observing cilia and flagella.

phase-contrast microscope

The ways in which organisms are grouped should reflect their evolution from common ancestors.

phylogenetic hierarchy

Used in step four of Gram stain procedure

safranin - Gram-positive cells remain purple, Gram-negative cells are pink

Type of electron microscope produces three-dimensional view of the surface of microbes and cellular structures whole species can be observed, but it only magnifies the external surface and requires a vacuum.

scanning electron microscope

Type of probe microscope that measures the flow of electrical current between the tip of a probe and the specimen to produce an image of the surface at atomic level.

scanning tunneling microscope

Type of stain used to determine size, shape, and arrangement of cells composed of a single basic dye they soak the smear in dye, rinse with H2O and blot dry

simple stains procedure

preparation of specimen techniques and why

smear, heat fixation, chemical fixation it is needed to attach microorganisms firmly on the slide and preserve their shape and size, so they aren't lost during staining

"A group of organisms that interbreed to produce viable offspring"

species

populations of cells that arose from a single cell - that share many stable properties, differ from others, and evolve as a group

strains

The science of classifying and naming organisms

taxonomy

bright field microscopes

the background is illuminated simple and compound microscopes may have a condenser lens

Type of electron microscope produces two-dimensional image of ultrastructure of cells on a fluorescent screen

transmission electron microscope

The ability to distinguish objects that are close together

resolution

Linnaeus proposed only two kingdoms: _____ and _____.

Animalia, plantae

Procedure that focuses on the separation on the basis the ability of microorganisms to use or produce certain chemicals (ferment carbohydrates) differences in fatty acid composition distinguish between bacteria rely on the ability of the organism to be cultured in the laboratory (pathogens such as Gram-negative Enterobacteria & Gram-positive bacteria, yeasts and fungi).

Biochemical tests

_____ microscopes have a greater _____ _____ and magnification than light microscopes and can magnify objects 10,000X to 100,000X.

Electron, resolving power

In endospore staining, why don't the spores stain red as well?

Heat from steam is used to drive the green primary stain into the endospores. Counterstaining is performed at room temperature, and the thick, impermeable walls of the endospores resist the counterstain

Type of differential stain that uses Gomori methenamine silver (GMS) and Hematoxylin and eosin (HE) stains to screen for the presence of fungi and the locations of carbohydrates in tissues as well as delineate many features of histological specimens, such as the presence of cancer cells.

Histological stain

What is different about Linnaeus taxonomic categories than the modern goal to distinguish organisms?

Linnaeus goal was to simply classify organism to catalogue them; the modern goal is to understand relationships among groups of organisms that reflect phylogenetic hierarchy and emphasis on comparison of organisms' genetic material.

Type of differential stain that highlights the presence of endospores produced by species in the genera Bacillus and Clostridium uses heat to drive the primary stain cool and rinse

Schaeffer-Fulton endospore stain procedure

Procedure that focuses on antibody-antigen interactions (antibody-antigen interactions are very specific) also utilizes the agglutination test that can be used to differentiate different species or even strains of the same species

Serology tests

Contains a single magnifying lens similar to magnifying glass used by Leeuwenhoek.

Simple microscope

_____ of microorganisms (thin film) made prior to staining.

Smear

Type of differential stain that distinguishes the genera Mycobacterium and Nocardia from other bacteria cover smear with tissue paper flood slide with stain and warm over steam remove tissue paper and cool rinse with HCl and alcohol counterstain

Ziehl-Neelson acid-fast stain procedure

Basic dyes stain _____ structures.

acidic structures by ionically noding to them and stain them works best in basic environments used more because most cells are negatively charged

Test in which an antiserum is mixed with a sample that potentially contains its target cells. In a positive result, clumps are formed; and in a negative result, there is no clumps.

agglutination test

empty magnification

an image that is magnified a lot but is faint and blurry

Procedure involving DNA sequencing and comparison determination of the organisms G+C ratio have provided a powerful tool for prokaryotic classification

analysis of nucleic acids

resolving power depends on calculated by

being able to distinguis between 2 objects the wavelength of the electromagnetic radiation the numerical aperture of the lens (the ability of a lens to gather light) resolution distance= .61 * (wavelength/numerical aperture)

Linnaeus used _____ _____ in his system of naming species

binomial nomenclature

Differences in intensity between two objects, or between an object and background.

contrast

Staining increases _____.

contrast

Use of light that is in phase increases _____.

contrast

Staining increases _____ and _____ by coloring specimens with stains/dyes.

contrast, resolution

Used in step one of Gram stain procedure

crystal violet - all cells are stained purple

Taxonomic key that contains a series of paired statements worded so that only one of two "either/or" choices applies to any particular organism; this key directs the user to another pair of statements, or provides name of organism.

dichotomous keys

Also called a Nomarski microscope create phase interference patterns and use prisms that split light beams into their component wavelengths (increases contrast and gives a 3D appearance) no staining required.

differential interference contrast microscope

Type of stain that uses more than one dye so that different cells, chemicals, or structures can be distinguished.

differential stains

Used in step three of Gram stain procedure

ethanol and acetone - smear is decolorized; Gram-positive cells remain purple, but Gram-negative cells are now colorless

Type of special stain where bacterial flagella become visible and allows determination of number and location of bacterial flagella

flagellar stain

Some cells are naturally _____, others must be stained.

fluorescent

Microscope that directs UV light source at specimen to radiate energy back as a visible wavelength (increases resolution and contrast due to shorter wavelength) used in immunofluorescence to identify pathogens and to make visible a variety of proteins.

fluorescent microscope

A convex lens focuses light on a _____ _____. The image is _____ and _____ as light rays pass the focal point and spread apart.

focal point, enlarged, inverted

Used in step two of Gram stain procedure

iodine - acts as a mordant; all cells remain purple

The use of light or electrons to magnify objects.

microscopy

In staining for electron microscopy, stains may bind to _____ in specimens or the _____.

molecules, background

Microscope that magnifies more than 100,000,000X and uses microscopic probes that move over the surface of a specimen.

probe microscopes

Magnification results when a beam of radiation _____ as it passes through a lens.

refracts

Oil immersion lens increases _____ because no light is refracted as it leaves the specimen and more light enters the lens. it also increases

resolution magnification

In fluorescent microscopes, UV light increases _____ and _____.

resolution, contrast