04 - Anaerobes, Uncultivatable Bacteria & Virology (MLS Exam #4) Flashcards
Define
Anaerobe
A bacterium able to replicate in the absence of oxygen.
Define
Aerotolerant anaerobe
Anaerobes that can survive a short exposure to oxygen but will not be able to perform metabolic processes unless placed into an anaerobic environment.
Define
Obligate or strict anaerobe
Anaerobes that are killed almost immediately in the presence of oxygen.
Define
Exogenous anaerobes
Anaerobes that exist outside the bodies of animals and the infections they cause are termed exogenous infections.
They can be found in soil, in freshwater and saltwater sediments,
Exogenous anaerobic infections are usually caused by gram-positive, spore-forming bacilli belonging to the genus Clostridium. Clostridia initiate infection when spores are ingested by way of contaminated food or gain access to the body through open wounds contaminated with soil.
Define
Endogenous anaerobes
Anaerobes that exist inside the bodies of animals and are the source of endogenous infections.
The anaerobes most frequently isolated from infectious processes in humans are those of endogenous origin.
Discuss the significance of anaerobes in the infectious process.
- Anaerobes outnumber aerobes on mucosal surfaces, such as the linings of the oral cavity, gastrointestinal (GI) tract, and genitourinary (GU) tract. These heavily colonized surfaces are the usual portals of entry into the tissues and bloodstream for endogenous anaerobes.
- Under ordinary circumstances, microorganisms that are members of the microbiota do not cause disease, and many actually can be beneficial.
- However, when some of these organisms gain access to usually sterile body sites, such as the bloodstream, brain, and lungs, they can cause serious or even fatal infections.
Define normal anaerobic flora for specimen types
Skin
Indigenous members of the skin microbiota include anaerobes that colonize the sebaceous glands and hair follicles.
Propionibacterium, peptostreptococci, Cutibacterium (Propionibacterium) acnes
Define normal anaerobic flora for specimen types
Intestinal tract
- It is estimated that 500 to 1000 different species of bacteria live in the GI tract. Microbiota studies have found that anaerobes outnumber facultative anaerobes by a factor of 1000 : 1.
- Any infection in the peritoneal cavity would likely be caused by organisms that have escaped from the GI tract.
Bifidobacterium, Eubacterium/ Eggerthella, Clostridium, peptostreptococci, Bacteroides fragilis group, Parabacteroides, Bilophila, Campylobacter, Fusobacterium, Porphyromonas, Prevotella, Sutterella, Veillonella
Define normal anaerobic flora for specimen types
Upper respiratory tract and Oral Cavity
Of the bacteria present in saliva, nasal washings, and gingival and tooth scrapings, 90% are anaerobes.
Actinomyces, Eubacterium/ Eggerthella, peptostreptococci, Campylobacter, Fusobacterium, Prevotella, Bifidobacterium, Porphyromonas, Veillonella
Define normal anaerobic flora for specimen types
Genitourinary tract
- Although anaerobic bacteria colonize the distal urethra, they are not considered to be a cause of uncomplicated urinary tract infections.
- Similarly, 50% of the bacteria in cervical and vaginal secretions are anaerobes.
- GU swabs and voided or catheterized urine specimens are unacceptable for anaerobic bacteriology because recovery of these organisms would not distinguish whether they were present as pathogens or as endogenous microbiota.
Peptostreptococci, Bifidobacterium, Fusobacterium, Lactobacillus, Mobiluncus, Prevotella, Veillonella
Discuss
Factors That Predispose Patients to Anaerobic Infections
Generally, infectious diseases involving anaerobic bacteria follow some type of trauma to protective barriers such as the skin and mucous membranes. Trauma at these sites allows anaerobes of the endogenous biota (or in some cases, soil anaerobes) to gain access to deeper tissues.
Human or animal bite wounds. Aspiration of oral contents into the lungs after vomiting. Tooth extraction, oral surgery, or traumatic puncture of the oral cavity. Gastrointestinal tract surgery or traumatic puncture of the bowel. Genital tract surgery or traumatic puncture of the genital tract. Introduction of soil into a wound.
Discuss the physical characteristics that are found when anaerobes are recovered in the infectious process.
*Infectious processes involving anaerobes are usually purulent, with many polymorphonuclear leukocytes present. (not always)
* Infection in close proximity to a mucosal surface
* Presence of foul odor
* Presence of large quantity of gas
* Presence of black color or brick red fluorescence
* Presence of sulfur granules
Many of the infectious processes involving anaerobes consist of mixtures of obligate anaerobes or mixtures of obligate or aerotolerant anaerobes and facultative organisms, this is called what?
Polymicrobial infection
Associate the causative anaerobic organism with the clinical infection.
Food poisoning
Clostridium perfringens
C. perfringens is associated with two types of food poisoning— type A, a relatively mild and self-limited GI illness, and type C, a more serious but rarely seen disease. C. perfringens foodborne disease usually follows the ingestion of large numbers of enterotoxin-producing strains in contaminated food.
Associate the causative anaerobic organism with the clinical infection.
Botulism
Clostridium botulinum
- Results from the ingestion of preformed botulinum toxin,Botulinum toxin is an extremely potent neurotoxin; only a small amount produces death.
- Weakness and paralysis are the main features of botulism.
- The food sources involved commonly in botulism include home-canned goods.
- Botulinum toxin type A (Botox) is also used medically to treat chronic migraines and temporarily improve facial wrinkles.
What is a common source of infant botulism?
Raw Honey
Associate the causative anaerobic organism with the clinical infection.
Tetanus
Clostridium tetani
- Tetanus occurs when spores in the environment enter the skin through puncture wounds.
- The clinical manifestations of tetanus are attributed to the neurotoxin tetanospasmin produced by Clostridium tetani. Tetanospasmin acts on neurons, preventing the release of inhibitory and excitatory neurotransmitters. This results in a spastic type of paralysis, with continuous muscular spasms. leading to trismus (lockjaw), risus sardonicus (distorted grin), and difficulty breathing.
- Therapy for tetanus requires the injection of antitoxin, muscle relaxants, and intensive therapy.
Associate the causative anaerobic organism with the clinical infection.
Myonecrosis (gas gangrene)
C. perfringens (the most common cause)
- Myonecrosis, or gas gangrene, usually occurs when organisms contaminate wounds, through trauma or surgery.
- C. perfringens, C. histolyticum, C. septicum, C. novyi, and C. bifermentans have all been associated with myonecrosis.
- C. perfringens is the most common cause. Under favorable conditions, the organisms are able to grow, multiply, and release potent exotoxins. In gas gangrene exotoxins, such as α-toxin produced by C. perfringens, cause necrosis of the tissue and allow deeper penetration by the organisms.
- The onset and spread of myonecrosis can be rapid, and extensive surgical debridement of the necrotic tissue is often required.** If treatment is delayed, amputation of the affected limb is not uncommon.**
Associate the causative anaerobic organism with the clinical infection.
Pseudomembraneous colitis
Clostridioides difficile
- This organism is part of the GI biota in about 5% of individuals
- Following antimicrobial therapy, many bowel biota organisms other than C. difficile are killed, thus allowing C. difficile to multiply with less competition and produce high levels of two toxins: toxin A, an enterotoxin, and toxin B, a cytotoxin.
- Bloody diarrhea with associated necrosis of colonic mucosa is seen in patients with pseudomembranous colitis. C. difficile is a common cause of health care– associated (nosocomial) infection.
Associate the causative anaerobic organism with the clinical infection.
Actinomycosis
Actinomyces spp. (A. israelii is the most common)
Actinomycosis is a chronic, granulomatous, infectious disease characterized by the development of sinus tracts and fistulae, which erupt to the surface and drain pus that may contain so-called sulfur granules, dense clumps of bacteria that may be colored.
Associate the causative anaerobic organism with the clinical infection.
Lemierre’s disease
Fusobacterium necrophorum
Lemierre disease, a syndrome of thrombophlebitis of the jugular vein that occurs rarely following group A streptococcal pharyngitis, is caused by F. necrophorum. The streptococcal infection produces a peritonsillar abscess containing a number of bacterial species. The abscess aids F. necrophorum in penetrating the tissue to reach the jugular veins.
Associate the causative anaerobic organism with the clinical infection.
Acne
Propionibacterium acnes
Which nonselective media should be used for anaerobic cultures?
Anaerobic blood agar (CDC)
An enriched medium containing sheep blood for enrichment and detection of hemolysis, vitamin K (required by some Porphyromonas spp.), and yeast extract
What selective media is used to select for Bacteroides fragilis group?
Bacteroides bile esculin agar (BBE)
- Containing gentamicin (which inhibits most aerobic organisms)
- 20% bile (which inhibits most anaerobes)
- Esculin; used primarily for rapid isolation and presumptive identification of members of the B. fragilis group
- Black colonies (because of esculin hydrolysis)
Name the selective medium containing sheep red blood cells and phenylethyl alcohol; used primarily to suppress the growth of any facultative, gram-negative bacilli (e.g., Enterobacteriaceae) that might be present in the clinical specimen, especially swarming Proteus spp.
Phenylethyl agar (PEA)
Supports growth of almost all obligate anaerobes (gram-positive and gram-negative)
What is the composition of anaerobic gas?
- 5% hydrogen
- 80-90% nitrogen
- 5-10% carbon dioxide (CO2)
Name 3 ways in which ANO2 conditions can be acheived?
- Anaerobic chambers
- Anaerobic jars
- Anaerobic bags
How long should primary anaerobic cultures incubate before an initial read?
48 hours
How long are anaerobic cultures routinely held for?
5-7 days (5 days at CCF)
* 10-14 to rule out Actinomyces or Cutibacterium acnes
When anaerobes are suspected, what steps must be performed and recorded for each colony morphotype present on the anaerobic blood agar plate to initiate presumptive identification of the isolates?
- Describe the colony morphology and note the media in which growth occurred. (CDC, BBE)
- Describe the Gram stain reaction and cell morphology.
- Set up an aerotolerance test
- Perform MALDI-TOF (if available)
Define
Aerotolerance Testing
Determines whether a microorganism isolated under anaerobic conditions is a strict or facultative anaerobe.
Subculture the suspected isolate to both aerobic (BAP) and anaerobic (CDC) media and observe the actual atmospheric requirements of the organism.
Interpretation of Aerotolerance Test Results
Describe the principle and application of the test.
Fluorescence
Porphyromonas and Prevotella fluoresce brick red under long-wave (366-nm) UV light. (Wood’s lamp)
Describe the principle and application of the test.
Catalase
Helps differentiates aerotolerant strains of Clostridium (catalase negative) from Bacillus (catalase positive).
15% hydrogen peroxide
Describe the principle and application of the test.
Spot indole test
The spot indole test is useful in identifying C. acnes (indole positive) from the similar Propionibacterium spp. (indole negative).
Describe the principle and application of the test.
Urease test
C. sordellii is the only Clostridium species that is urease positive.
Describe the principle and application of the test.
Lipase production (EYA)
**Lipase-positive organisms produce a colony covered with an iridescent, multicolored sheen, sometimes described as resembling the appearance of gasoline on water or mother of pearl.
Fusobacterium necrophorum
Describe the principle and application of the test.
Lecithinase production (EYA)
Lecithinase cleaves lecithin found in egg yolk, releasing insoluble fat (diglyceride) that produces an opaque zone around the colony.
C. perfringens
Describe the principle and application of the test.
Sodium Polyanethol Sulfonate Disk (SPS)
(SPS) disk aids in the identification of anaerobic gram-positive cocci.
An SPS-sensitive, gram-positive anaerobic coccus can be presumptively identified as
Peptostreptococcus anaerobius
An SPS-resistant, spot indole– positive, gram-positive anaerobic coccus can be presumptively identified as
Peptoniphilus asaccharolyticus
Name 2 rapid anaerobic identification panels.
- RapID ANA
- API 20 A
Discuss antimicrobial susceptibility testing considerations for anaerobic bacteria.
Beacuse the isolation, identification, and susceptibility testing of anaerobes is slow, when physicians suspect an anaerobic infection, they routinely select a broad-spectrum agent for empiric therapy that will cover most anaerobes, pending outcome of the culture.
Carbapenems, metronidazole, and the β-lactam combination antibiotics continue to have activity against most anaerobes and are used as empiric therapy.
When should susceptibility testing be performed on anaerobes?
When it is recovered as a pure culture in a sterile site.
- Brain abscess * Endocarditis * Infection of a prosthetic device or vascular graft * Joint infection * Osteomyelitis * Bacteremia
A test for beta-lactamase production VIA cefinase is disc is performed on what type of anaerobic organisms?
Anaerobic GNB (other than B. fragilis group)
What method(s) are used to perform anaerobic susceptibility testing?
- E-test
- Microbroth dilution panels
Other than susceptibilty testing, how can anaerobe-associated diseases be treated?
- Surgery
- Hyperbaric oxygen therapy
- Antitoxins
- Refractory CDAD can be treated with fecal microbiota transplant
What are some indications that anaerobes may be present in a Culture?
- A foul odor on opening an anaerobic jar or bag.
- Colonies present on the anaerobically incubated blood agar plates but not on the CO2-incubated blood plate
- Good growth (> 1 mm in diameter) of black colonies on a BBE agar plate, characteristic of members of the B. fragilis group
- Double zone of hemolysis on CDC (ANO2) suggestive of C. perfringens
Presumptively Identify the Gram-Positive Anaerobes
- Colony morphology: Large, flat colonies; barnyard odor, chartreuse fluorescence
- Gram stain: Thin rods, rare spores
Clostridium difficile
Presumptively Identify the Gram-Positive Anaerobes
- Colony morphology: Large, irregular-shaped, double zone of β-hemolysis
- Gram stain: Boxcar, large, square rods
Clostridium perfringens
Presumptively Identify the Gram-Positive Anaerobes
Colony morphology: Smoothly swarming
Gram stain: Thin rods, subterminal spores
Clostridium septicum
Is often a marker for gastrointestinal cancer
Presumptively Identify the Gram-Positive Anaerobes
Colony morphology: Smoothly swarming but slow growing
Gram stain: Swollen terminal spores
Spot Idole: Positive
Clostridium tetani
Which anaerobes are most commonly associated with a gram stain of gram positive cocci?
Peptostreptococcus spp., Finegoldia, or Peptoniphilus
What anaerobe has a gram stain of Gram negative cocci?
Viellonella spp.
What anaerobe has a gram stain of “fusiform” (thin with pointed ends) GNB and colonies that resemble “breadcrumbs”?
Fusobacterium nucleatum
What anaerobic species has a gram stain of branching, filamentous, non-spore forms gram positive rods?
Actinomyces spp.
Which species of Actinomyces form “molar tooth” colonies?
Actinomyces israelii
Describe Actinomycosis
Actinomycosis is a chronic, granulomatous, infectious disease characterized by the development of sinus tracts and fistulae, which erupt to the surface and drain pus that may contain so-called sulfur granules, dense clumps of bacteria that may be colored.
Actinomyces has a similar gram stain to Nocardia. What stain could be used to differentiate the two?
AFB stain. Nocardia is Acid fast postive. Actinomyces is acid fast negative.
Which anaerobe has a gram stain of non-spore forming “spidery” gram positive bacilli and the colonies described as white and “puffy”?
Cutibacterium acnes
What reactions does Cutibacterium acnes have to spot indole and catalase?
Positive for both
Cutibacterium acnes is a slow growing organism, typically incubated for 10-14 days, and is associated with what type of infection?
Prosthetic joint infection
What organism is associated with IUD (intrauterine device) infections?
Actinomyces
How long should a culture be incubated for when trying to rule out Actinomyces or Cutibacterium acnes?
10-14 days
What anaerobe has a gram stain of pleomorphic GPB that looks like a “dog biscuit”?
Bifidobacterium
Which anaerobic organism may be considered a contaminant if isolated from a blood culture bottle?
Cutibacterium acnes
It can be part of skin flora
Which anaerobic species is associated with spore forming GPB?
Clostridium spp
What is the gram stain of Bacteroides spp?
Bacteroides spp. colonies reveal gram-negative coccobacilli or bacilli.
To be diagnosed with C. diff colitis, you must test positive for what?
Toxins
Describe
Spirochetes
Slender, flexuous, helically shaped, unicellular bacteria ranging in size from 0.1 to 0.5 µm wide and from 5 to 20 µm long, with one or more complete turns in the helix.
How do humans contract disease by leptospires?
Through contact with the urine of an infected animal. The usual portal of entry being conjuctiva or abrasions or cuts in the skin.
Leptospirosis is also known as what disease?
Weil disease
Severe systemic disease includes renal failure, hepatic failure, and intravascular disease and can result in death.
What media is used to isolate leptospires?
Fletcher’s media
What organism is the causitive agent of Lyme disease?
Borrelia burdorferi
What vector transmits Lyme disease?
Ticks known as Ixodes
Explain the three stages of Lyme disease.
Stage 1: Erythema migrans (localized circular rash at innoculation site)
Stage 2: Neurologic or cardiac involvement: most commonly facial nerve palsy
Stage 3: Joints most commonly involved (arthritis)
Describe the diagnostic method used to diagnose Lyme disease
IFA or EIA and confirmation with Western blot
Name the disease associated with Borrelia recurrentis
Relapsing fever.
Fevers that subside after 3-6 days and then return in 1 week
What vector is associated with endemic relapsing fevers?
Ticks
What vector is associated with epidemic relapsing fevers?
Louseborne
Lice
Louseborne fever is transmitted via the body louse, Pediculus humanus, and humans are the only reservoir.
What is the causative agent of syphilis?
Treponema pallidum
Describe the microscopy used to visualize T. pallidum in clinical specimens.
The organisms are too thin to be observed by bright-field microscopy, so spirochetes are illuminated against a dark background. Dark-field microscopy requires considerable skill and experience; however, demonstration of motile treponemes in material from the chancre is diagnostic for primary syphilis.
Describe the 3 stages of the disease caused by T. pallidum.
- Primary: A chancre or ulcer appears at site of innoculation and lasts 1-8 weeks
- Secondary: Flulike symptoms, swollen lymph nodes and rash
- Tertiary or late stage: 1/3 of untreated patients progress to 3rd stage; lesions (gummas) seen in bone, skin & other tissues.; neuro & cardiovascular involvement
Name the drug of choice for treating patients with syphilis
Penicillin
The Life cycle of Chlamydia.
Chlamydia’s life cycle has a unique growth cycle that involves two distinct forms, an elementary body (EB), and a reticulate body (RB), which form is infectious?
The elementary body (EB) is infectious, and the reticulate body (RB) is noninfectious.
C. trachomatis causes what type of eye infection?
C. trachomatis causes the chronic eye infection trachoma, the number one cause of preventable blindness in the world.
Trachoma is a chronic disease that begins as follicular conjunctivitis. The chronic inflammation causes the eyelid to turn inward, which results in continual abrasion to the cornea from the eye lashes. The condition results in scarring and ulceration of the cornea. This can result in secondary bacterial infection and blindness.
Name the organism that is an obligate intracellular parasite that infects the non-ciliated, columnar or transitional epithelial cells of the conjunctiva, respiratory, UG tract or rectum.
Chlamydia spp.
Chlamydia is usually susceptible to what drugs?
Doxycycline
What is the most common STD in the United States whose infection is often asymptomatic?
Chlamydia trachomatis
Diseases associated with C. trachomatis.
Associated with eye trachoma, lymphogranuloma venereum (LGV), PID, and Reiter syndrome.
How is the Chlamydia detected?
Nucleic Acid Hybridization and Amplification Assays.
NAATs
- While cell culture is the gold standard: it has been replaced by NAATs because of it’s higher sensitivity and specificity.
- Can be performed on urine and urethral/cervical swabs.
- Can detect two STDs in one sample— gonorrhea and C. trachomatis infection.
- Technicially less demanding.
- However, as of yet no NAAT has been approved for use on conjunctival, oropharyngeal, or rectal specimens.
What strains of C. trachomatis are more invasive, producing a more serious infection and pronounced antibody response?
LGV Strains
What type of species of Chlamydia is recognized as an important respiratory pathogen?
Chlamydophila pneumoniae
formally known as TWAR
Respiratory pathogen known to be the cause of sinusitis, pharyngitis, acute respiratory disease, bronchitis and pneumonia. Implicated as a possible factor in asthma and cardiovascular disease. Resembles that of Mycoplasma pneumoniae
Which organism causes psittacosis, a contagious disease of birds that is transmissible (especially from parrots) to human beings as a form of pneumonia?
Chlamydophila psittaci
Name the organism associated with the Rickettsial disease:
Rocky Mountain Spotted Fever (RMSF) and the vector is a dog tick.
R. rickettsii
Most severe of the Rickettsial infections. Acquired through tick bites (Dermacentor variabilis) in the USA – the bacteria then enter the blood stream and are phagocytosed where replication can occur intracellularly. Symptoms occur after 7 days of incubation – Rash (patches on wrists and ankles), fever, headache, nausea, etc. High mortality rate without treatment
Name the organism associated with the Rickettsial disease:
Murine typhus and the vector is a flea.
R. typhi
Humans scratch the bite of the flea, pushing flea stool into the wound, which contains the bacteria.
Name the organism associated with the Rickettsial disease:
Epidemic typhus and the vector is a louse (Pediculus humanus).
R. prowazekii
. Humans scratch the bite of the lice, pushing the lice stool into the wound, which contains the bacteria. Rash of the palms and soles of the feet occurs. Mortality in untreated patients can reach 40%.
Name the organism associated with the Rickettsial disease:
Scrub typhus and the vector is a chigger.
Orientia
Name the organism associated with the Rickettsial disease:
Rickettsialpox, the reservoir is the common house mouse, and the vector is the mouse mite Liponyssoides sanguineus.
R. akari
Name the organism associated with the Rickettsial disease:
Human granulocytic anaplasmosis (HGA) and the vector is a deer tick.
Anaplasma phagocytophilum
Note that As the cells divide within the phagosome, they develop into morulae (mulberry-like bodies; Fig. 24.11). Morulae are round to oval clusters of bacteria 1 to 3 µm in diameter. As the host cell ruptures, the morulae break into many individual EBs, which continue the infective cycle.
Name the organism associated with the Rickettsial disease:
Human Granulocytic Ehrlichiosis (HME) and the vector is the lone star tick.
Erlichia
Name the organism:
Causative agent of Q fever and can infect a wide range of vectors (ticks, birds, fish, livestock and other mammals).
Coxiella burnetii
- Considered to be a facultative intracellular parasite.
- Spore-former.
- Highly contagious and considered a bioterrorism agent.
- IFA of infected tissue is a reliable identification method
Name the organism:
Cat scratch fever and the vector is a kitten.
Bartonella henselae
True or False: Chlamydiae and rickettsiae are both obligate intracellular organisms.
True
What organisms should be considered as possible causes of neonatal conjunctivitis?
N. gonorrhoeae and C. trachomatis
What organisms are characterized by being the smallest free-living bacteria and lack a cell wall?
Mollicutes or Mycoplasma
Mycoplasma and Ureaplasma are the two genera in the family Mycoplasmataceae.
Mycoplasma pneumoniae causes which disease?
Primary atypical pneumonia (walking pneumonia)
Mycoplasma hominis is associated with what disease?
Urogenital tract disease
Ureaplasma urealyticum is associated with what disease?
Urogenital tract disease
Why can’t you gram stain Mycoplasma?
Because they lack a cell wall and do not retain the stain
Often isolated on anaerobic media and after performing a gram stain on the tiny colony with a result of “no organisms seen”, Mycoplasma should be suspected
Mycoplasmas are generally slow-growing, highly fastidious, facultative anaerobes requiring complex media containing what for growth?
Cholesterol and fatty acids
Which species of Mycoplasma form colonies with slightly raised centers, giving the classic “fried egg” appearance?
M. hominis
Describe the structural components of viruses
At a minimum, viruses contain a viral genome of ribonucleic acid (RNA) or deoxyribonucleic acid (DNA) and a protein coat— the capsid. The genome can be double stranded (ds) or single stranded( ss).
The genome and its protein coat together are referred to as what?
The nucleocapsid
The entire virus particle is called what?
Virion
How are viruses classified?
- Genome type (RNA or DNA)
- Number of strands in the genome (ds or ss),
- Morphology
- Presence or absence of an envelope
Viruses are obligate intracellular parasites. What does that mean?
They must be inside a living cell and use the host cell machinery to replicate.
Describe the basic steps of viral replication
- Absorption. In the first step for infection of a cell to occur, virions must absorb or attach to the cell surface.
- Penetration. Viruses can penetrate the cell by several different mechanisms and penetration is virus dependent.
- Uncoating.Once inside the cell, the virus loses its protein coat, releasing the genome. The viral genome then directs the host cell to make viral proteins and replicate the viral genome.
- Assembly or maturation of the virus particles.The new virions are formed and then released by lysis if they are naked viruses or by budding if they are enveloped viruses.
When should samples be collected for viral detection?
Viral shedding is highest in** early stages of infection** and decreases rapidly a few days after acute symptoms.
What is the optimal specimen for viral detection?
For optimal recovery, specimens for viral isolation should be **collected from the affected site. **
For example, secretions from the respiratory mucosa are most appropriate for viral diagnosis of respiratory infections. Aspirates, or surface swabs, are usually appropriate for lesions.
What types of swabs can and cannot be used for viral specimen collection?
- Swabs used must be made of Dacron or rayon.
- Do not use Calcium alginate or wood shafted swabs. They can inhibit the replication of some viruses and can interfere with nucleic acid amplification tests.
If a viral specimen must be frozen, what temperature should it be kept at?
–70 ° C
–70 ° C. Specimens should never be stored at − 20 ° C because this temperature facilitates the formation of ice crystals that will disrupt the host cells and result in loss of viral viability. Repeated freeze– thawing cycles are to be avoided because they can also result in loss of viral viability.
Define cytopathic effect (CPE) as it relates to viral identification
Many viruses produce distinctive and characteristic visual changes in infected cells referred to as a cytopathic effect (CPE).
Some viruses produce a very characteristic CPE that can provide a presumptive identification of a virus isolated from a clinical specimen. For example, HSV grows rapidly on many different cell lines and frequently produces a CPE within 24 hours. A predominantly cell-associated virus, HSV produces a focal CPE (in which adjacent cells become infected) and plaques, or clusters of infected cells. The combination of rapid growth, plaque formation, and growth on many different cell types is presumptive evidence for the identification of HSV.
Name 4 general types of cell cultures / lines used for viral isolation
- HDF, human diploid fibroblasts
- HEp2, human laryngeal carcinoma cell line
- PMK, primary monkey kidney
- RK, rabbit kidney
Describe the four major methods the clinical laboratory uses to diagnose viral infections.
- Direct detection of the virus in clinical specimens (microscopy, EIA)
- Nucleic acid– based detection (PCR)
- Isolation of viruses in cell cultures (gold standard but seldom used)
- Serologic assays to detect antibodies to virus
Name the phenomenon where slight antigenic change is seen in influenza viruses over time because of minor mutations in the ssRNA.
Antigenic drift
Name the phenomenon whereby an often-unexpected change occurs in influenza virus strains. This antigenic change is often so drastic that it triggers pandemics. Causes changes in H and N antigens.
Antigenic shift
Name that Virus
Most cases are asymptomatic. Those that are symptomatic usually involve the respiratory tract, eye, or gastrointestinal tract.
Adenovirus
dsDNA
Family: Adenoviradae
Name that Virus
There are two species: HSV-1 and HSV-2. These viruses cause oral herpes, genital herpes, neonatal herpes, and ocular herpes. Also the leading cause of fatal sporadic encephalitis in the United States. Neonates are most at risk for HSV-2 encephalitis. HSV detection by culture is sensitive and is often positive in less than two days.
Herpes simplex viruses
(aka Human herpesvirus 1 and Human herpesvirus 2)
dsDNA
Family: Herpesviridae
Name that Virus
Congenital infection can be significant. If the mother acquires primary infection during pregnancy irreversible damage could result to the fetus. As a result, this virus is one of the leading causes of mental retardation, deafness, and intellectual impairment. Infants may develop a blueberry muffin rash.
Cytomegalovirus
(Human herpesvirus 5)
dsDNA
Family: Herpesviridae
Name that Virus
Virus infects B lymphocytes and stimulates the production of heterophile antibodies. Infection can present as infectious mononucleosis. Some cancers have been associated with this virus including Burkitt’s lymphoma. Atypical lymphocytes may been seen in hematological testing.
Epstein-Barr virus
(Human herpesvirus 4)
dsDNA
Family: Herpesviridae “kissing disease”
Name that Virus
Causative agent of chickenpox and shingles. Skin scrapings are the best specimens.
Varicella-Zoster virus
(human herpesvirus 3)
dsDNA
Family: Herpesviridae
Name that Virus
Associated with the childhood disease called roseola infantum.
Human herpesvirus 6 (HHV-6)
dsDNA
Family: Herpesviridae
Name that Virus
Associated with genital warts. Diagnosis is made from koilocytes on pap smears. Linked to cervical cancer.
Human papilloma virus (HPV)
dsDNA
Family: Papillomaviridae
Name that Virus
Infections are associated with Kaposi sarcoma.
Human herpesvirus 8 (HHV-8)
Name that Virus
Causative agent of smallpox.
Variola virus
dsDNA
Family: Poxviridae
Name that Virus
The most recognized symptom of an infection caused by this virus is erythema infectiosum (also called fifth disease) - a rash on the cheeks. Some referred to it as a slapped cheek appearance.
Parvovirus B19
ssDNA
Family: Parvoviridae
Name that Virus
Most common cause of viral gastroenteritis in infants and children.
Rotavirus
dsRNA
Family: Reoviridae
Name that Virus
A group of viruses transmitted by arthropods such as ticks and mosquitoes and can cause hemorrhagic fevers in humans.
Arboviruses
Name that Virus
A group of viruses carried by rodents and can cause hemorrhagic fevers in humans
Arenaviruses
Name that Virus
Causes gastroenteritis and is associated with outbreaks on cruise ships.
Norovirus
Name that Virus
Novel strains of this group of viruses has been associated with severe acute respiratory syndrome (SARS).
Coronaviruses
ssRNA
Family: Coronaviridae
Name that Virus
Infections with these two viruses are rare, but almost always fatal due to hemorrhagic fevers.
Ebola and Marburg virus
ssRNA
Family: Filoviridae
Name that Virus
These viruses have a segmented genome and are classified based on two major surface glycoproteins: hemagglutinin (H) and neuraminidase (N). They attack ciliated epithelial cells lining the respiratory tract and cause the flu.
Influenza virus A, B, and C
ssRNA
Family: Orthomyxoviridae
Name that Virus
These viruses are the primary cause of respiratory disease in young children. Type 1 is the primary cause of croup.
Parainfluenza viruses (1 through 4)
ssRNA
Family: Paramyxoviridae
Name that Virus
ssCauses bilateral swelling of the parotid glands.
Mumps virus
ssRNA
Family: Paramyxoviridae
Name that Virus
This virus causes a stuffy nose, cough, and Koplik spots.
Measles virus (also known as rubeola)
ssRNA Family:Paramyxoviridae
Koplik spots are white spots that appear on the mucous membranes of measles patients approximately 1 day before the appearance of the typical measles rash.
Name that Virus
Most common cause of severe lower respiratory tract disease among infants and young children.
Respiratory syncytial virus (RSV)
ssRNA
Family: Paramyxoviridae
Name that Virus
Infection with this virus can be assumed if a child has a lower respiratory tract infection, but tests negative for respiratory syncytial virus, influenza viruses, and parainfluenza viruses.
Human metapneumovirus
ssRNA
Family: Paramyxoviridae
Name that Virus
Causes nausea and vomiting in neonates.
Enterovirus
Name that Virus
Virus requires an RNA-dependent DNA polymerase to replicate and is T-lymphotropic (infects CD4+ T-cells). Patients are treated with highly active antiretroviral therapy (HAART).
HIV (human immunodeficiency virus)
Name that Virus
Infection usually results after being bitten or scratch by a wild animal. Symptoms are flulike at first and then develop into mental status changes. Patients often experience hallucinations, paralysis, excessive salvation, hydrophobia, and bouts of terror.
Rabies virus
Name that Virus
These viruses are a diverse collection of viruses grouped together because they all infect primarily the liver. Laboratory diagnosis is based on serologic markers.
Hepatitis A, B ,C, D,E, etc.
Name that Virus
German measles
Rubella virus
Name that Virus
The common cold
Rhinovirus
Name that Virus
Infects the central nervous system and cause paralysis.
Polioviruses
Name that Virus
Hand/foot/mouth disease: Small painful sores appear on the tongue, buccal mucosa, and soft palate and a rash on the hands, feet, and buttock.
Cosackievirus A
How do retroviruses (e.g., HIV) replicate?
Retroviruses replicate with the enzyme reverse transcriptase, which uses viral RNA as a template to make a complementary DNA strand.
Members of which family produce life-long latent infections?
Herpesviridae
Which opportunistic infections or conditions are used as indicators of acquired immunodeficiency syndrome (AIDS)?
Some opportunistic infections and conditions associated with AIDS include candidiasis, cryptococcal meningitis, cryptosporidiosis, histoplasmosis, persistent HSV infections, mycobacterial infections, recurrent pneumonia, and Kaposi sarcoma.
Which immunologic markers are used to diagnose human immunodeficiency virus (HIV) infection?
Testing for HIV-specific antigens and antibodies is important in the diagnosis of HIV infection, including antibodies to viral antigens p24, p31, gp41, and gp120/ 160.
Why are vaccines for influenza not always effective?
Influenza viruses mutate often as a result of replication errors. These mutations cause antigenic drift, ensuring antigenic variability of strains each year. Recombination events of the influenza A genome result in a major antigenic change called antigenic shift. Health care agencies predict the most likely strains that will predominate in the next season. Trivalent influenza vaccines are available prior to the start of the influenza season. Although this process is extremely successful, occasionally an unexpected strain will predominate, and the vaccine may not provide total coverage for that strain. Even with this known possibility, it is still advised that all persons be vaccinated because some protection is better than no protection at all.
