Case Study 1 Perspectives
Plankton is the assemblage of living organisms floating with currents in upper regions of aquatic
environments such as lakes and oceans. In addition to the bacteria (called bacterioplankton)
targeted by the study, plankton contains microscopic algae, which are termed phytoplankton, and
protozoa and tiny animals, which make up the zooplankton. One would also expect to find some
archaea and viruses. Microscopic fungi such as molds and yeasts are less likely to occur in the
planktonic community.
Other specialists that could be involved in further studies of the plankton are marine
ecologists, molecular biologists, biochemists, geomicrobiologists, taxonomists, bacteriologists,
algologists, protozoologists, virologists, geneticists, and microbiologists with expertise in specific
groups or species of microorganisms.
The ocean makes up three-fourths of the earth’s surface area, and it extends down several
miles into a giant basin carved out of the earth’s crust. The existence of vast areas that have yet to
be searched and sampled means that many unknown life forms are yet to be discovered. Very
recently microbiologists identified species of amoebas living in the ocean that harbored the
largest viruses ever discovered. These unusual viruses have a structure that is not typical of
known viruses and yet they are not cells. A number of virologists are postulating that they could
constitute a new, fourth domain. It will take hundreds of years of scientific exploration to reveal
all of the hidden secrets of planet earth.
This ambitious undertaking was just the beginning. It was followed by additional
voyages by Sorcerer II and a team from the Marine Biological Institute in Woods Hole,
Massachusetts, and it is continuing today all over the globe. Dr. Venter plans to use his findings
to explore potential applications for microorganisms, including developing synthetic bacteria for
creating new chemical products, bioremediation of wastes, and developing alternative forms of
energy.
Case Study 2 Perspectives
Water is composed of two hydrogens covalently bonded to a single oxygen. The nature of the
bonding makes water molecules polar, that is, having ends with opposite charges. In its liquid
form, its polarity makes it an excellent solvent for dissolving ions and numerous biological
compounds. Cells require molecules dissolved in solution for most chemical reactions, which are
involved in membrane function, digestion, synthesis, and other metabolic activities.
It is now clear that Mars has abundant water, but mainly in the ice and vapor form. A
major unknown is whether any significant liquid water currently exists. There are surface
features that point to a past history of liquid water, but right now it appears that the ice converts
directly into water vapor when exposed to increasing temperatures. Some locations on the planet
may have “hot spots” deeper in the crust where liquid water exists part of the time. It is here that
any remnants of life are most likely to occur.
Carbon is a versatile element that can make four bonds with numerous other atoms,
including other carbon atoms. It forms compounds with elongate chains, side chains, and rings
that make it possible to construct the complex macromolecules like protein and DNA that are so
crucial to functions and structures in cells.
It is clear that inorganic carbon in the form of carbon dioxide occur in places in the solar
system that could not support life. So, even though inorganic carbon may be involved in cellular
reactions like photosynthesis and respiration, non-living processes may also create them. With
organic compounds, something as simple as methane may also be produced by non-living
reactions, and it is not a firm indicator of life. More complex compounds such as sugars and
amino acids tend to be associated with life functions and are more reliable evidence. The
discovery of intact proteins or nucleic acids would be a chemical signature that truly points to
life. These complex macromolecules must be synthesized, and would not ordinarily arise by
random chemical reactions.
Other elements that would ordinarily be involved in the chemistry of biological forms are
oxygen gas, phosphorus, sulfur, nitrogen gas, magnesium, iron (iron oxide is the reason Mars has
a red color), sodium, potassium, calcium, and chloride. All of these have been tested for and
found to be present in the atmosphere or soil. So, chemically at least, the major participants for
life as we know it exist on Mars.
Case Study 3 Perspectives
Signs and symptoms are noticeable manifestations in a patient that can help direct
diagnosis of a disease and help pinpoint which anatomical sites are affected. A sign is an
objective observation that is measurable, like a fever or white blood cell count. A symptom is a
subjective report related to something a patient feels, for instance, a headache or pain. The most
important initial diagnostic signs of infection in this case are an elevated white blood cell count,
cloudy spinal fluid, and splotches on the legs. The most important symptoms are headache, stiff
neck, and mental confusion.
Monitoring the cerebrospinal fluid or CSF provides a means to quickly assess the
presence of infectious agents in the spinal column and the brain. The CSF bathes the brain, spinal
cord and membranes and is ordinarily completely free of microbes, or sterile. If it is cloudy
macroscopically, this could indicate growth of bacteria or other infectious agents. A microscopic
inspection can provide immediate feedback as to a possible cause of the cloudy CSF.
A Gram stain is one of the key tests for getting quick feedback on the kind of microbes
that might be present in a sample. It is routine in meningitis because it can differentiate among
several types of bacteria and detect certain other infectious agents, but it will not detect viruses.
Within moments, a lab technician can tell if there are bacterial cells, and they can identify their
gram reaction and shape. For example, in this case the result showed gram-negative cocci, but
meningitis is also commonly caused by Streptococcus pneumoniae, which would stain as grampositive cocci, and Haemophilus influenzae, which would appear as gram-negative rods. An early
clue to the microorganism also helps to instruct the types of drugs that will be given. Since
meningitis can cause death in 5% to 10% of people in a few hours, early drug treatment is critical.
Neisseria meningitidis is routinely isolated and grown on blood agar, chocolate agar, and
Thayer Martin medium. Its identity can be confirmed by a series of biochemical tests that
differentiate it from close relatives that may look microscopically similar. See figure 3.13 for an
example.
Case Study 4 Perspectives
All of the symptoms and signs in this case can be linked to endocarditis, but most of them
are too non-specific to be helpful in diagnosis. The most important considerations are the constant
fever, the history of a replacement heart valve, and the respiratory and circulatory symptoms. A
fever of unknown origin (FUO) is often traced to a chronic infection. The artificial heart valve
provides an opportunity for microorganisms to grow in the body because it readily supports a
biofilm that will not be protected by the immune system. The dysfunctional valve causes blood
to back up in the heart, leading to respiratory distress and abnormal heart function.
The reason that Mr. Jones throat cultures were negative was because his throat was not
the source of the pathogen. It was a blood-borne infection, also known as septicemia. The patient
might have survived if the attending physician had known about the heart valve and a prior
infection early on. If he had taken blood cultures earlier, the infectious agent could have been
isolated and identified and the correct treatment started.
Staphylococcus aureus is the most common cause of endocarditis, especially associated
with artificial heart valves. MRSA stands for methicillin-resistant Staphylococcus aureus, which
is a shorthand way of saying that the microbe is resistant to numerous drugs used in treatment.
The reason that the penicillin Mr. Jones was given to take at home would not have worked is that
most strains of S. aureus are resistant to it. Treatment with a mixture of drugs is indicated, but it
needs to start early in the infection. MRSA is an emerging pathogen that started as a problem in
the hospital, but is now prominent in non-hospital settings as well.
Case Study 5 Perspectives
Parasitology is the study of eukaryotic organisms called parasites that invade the body
and cause disease. It includes two major groups: the protozoans--primarily single-celled
microorganisms, and the helminth worms-- multicellular invertebrate animals that may or may
not be microscopic. Parasitologists study the morphology, life cycles, epidemiology, and
pathology of these parasites. A list of the most common parasitic diseases in humans would total
around 35, distributed all around the globe.
Dr. Hotez’ quote about the estimated worldwide case rate is meant to emphasize the
enormity of the situation. In those regions where neglected tropical parasites prevail, the
populations could be suffering from several different NTDs simultaneously, so the actual disease
burden could easily top 3 or 4 billion cases overall.
A vector is any animal that passes an infectious agent to humans. Vectors may be
invertebrates or vertebrates. The most common vectors are arthropods such as insects
(mosquitoes, fleas, bugs, biting flies) and arachnids (ticks and mites). Vectors of this type usually
have a unique relationship with their parasite, serving as an alternate host that supports a different
phase of development for the vector. Many vectors infect humans by taking a blood meal,
thereby providing a route of entry for the parasite.
Dr. Hotez has been deeply involved in educating about and finding solutions for the
terrible legacy of these diseases of poverty. He has partnered with the Gates Foundation to
develop a vaccine for hookworm and written dozens of articles and a book on the subject.
Recently Global Network in partnership with END7 has developed programs to oversee the world
funding for 7 of these diseases with the goal of eradicating them by the year 2020. These
organizations support the “Just 50 cents” campaign, which advocates that a donation of only 50
cents will provide the necessary medication to save a child’s life.
Case Study 6 Perspectives
Viruses must attach to their host cells by their receptor molecules before they can enter
the cells and complete their infection cycle. The shape of these receptors is complementary to
specific molecules on the host cell. This specificity of attachment limits the kinds of cells they
can invade. Once they are bound, viruses go to the next phase, which is their entry into the
interior of the cell and production of new viruses. The flu virus attaches by its H receptors to the
epithelial cells of the upper respiratory tract. From here it is poised to fuse with the cell and
continue its path into the cell. N receptors have the important functions of helping the virus to
penetrate the respiratory mucosa and facilitating the release of new viruses from infected cells.
Even though influenza virus has host specificity, infection does not require that all of its
receptors be specific to its primary host. For example, a swine virus could have half of its
hemagglutinin receptors specific to swine, but the other half could be specific to humans. As a
result it can infect both species. The same would be true with bird influenza viruses. The
underlying mechanism comes down to viral genetics. If a virus has developed mutated genes that
give rise to receptors that fit another host, then that other host will be susceptible to infection.
A pandemic is generally defined as an epidemic that spreads across continents. Health
officials consider influenza at pandemic levels when a new virus emerges that many people are
susceptible to, when there is sustained human to human transmission, and when infection
becomes widespread globally. The rapid spread of this pandemic can be attributed to factors such
as its prevalence and rate of travel, the ready spread of the virus through the air by means of
respiratory secretions, and the lack of prior exposure and immunity in a large number of people.
One benefit of prior influenza infections is that a person usually develops long term
immunity to that strain of virus. Older members of the population are likely to have encountered
viruses in the past that were similar enough to 2009 H1N1 that protective immunities from the
earlier infection also worked to counteract the new virus. Thus a person with prior immunities
will be able to inactivate the virus before it can get a foothold. This is the same idea behind the
flu vaccines--they prevent the infection by getting the immune system ready to respond to the
virus in the future.
Case Study 7 Perspectives
Extremophiles that can survive the toxic conditions of the pit are represented by all
microbial groups. In all, 130 unique species of microorganisms have been identified so far. They
include prokaryotes—the archaea and bacteria, which are best known for their capacity to adapt
to extremes, but algae, protozoa, and fungi have also been found in abundance. Though it is true
that animals and plants cannot adapt rapidly to such toxic conditions, many types of single celled
microorganisms can alter their metabolism in a relatively short time; this coupled with their short
generation times—hours vs years—they are much better at adapting to extreme conditions.
In being photosynthetic, the algae are a prolific source of oxygen gas that promotes
survival and growth of aerobic organisms--namely, fungi, protozoa, and many types of bacteria.
In addition, algae play an important role in the biogeochemical cycling of several elements-nitrogen, phosphorus, sulfur, and carbon--that all organisms use in metabolism. Algae also add
carbonates to the water which in time could buffer the pH and reduce its acidity.
Bioremediation is the introduction of microbes into polluted environments to reduce or
degrade the toxic chemicals present there. Some microbes absorb heavy metals and others help to
precipitate them into insoluble compounds which could be more readily removed. Since the
organisms that naturally detoxify the pit ecosystem seem to be already present, a viable solution
would be to enrich the water with extra inorganic nutrients such as nitrates and phosphates that
favor algal growth. It is expected that the increased population of algae would absorb the toxic
metals and concentrate them in the sediments at the bottom of the pit, where they would be less
accessible and less likely to leak out.
Case Study 8 Perspectives
Biochemically, fermentation is the partial breakdown of simple carbohydrates that does not involve oxygen
as a final electron acceptor, and that releases a wide variety of products along with a small amount of
energy (ATP). The term is also used collectively in a later chapter (27) to define any large scale industrial
processes for synthesizing commercial products such as drugs, foods, and other organic chemicals.
Fermentations begin with a usable substrate such as simple sugars (glucose), amino acids, or other
small organic compounds. The process requires microbes with the necessary metabolic pathways and
enzymes to carry out a particular type of fermentation. Fermentation proceeds under anaerobic conditions
in a medium that contains additional essential nutrients for growth (all major bioelements) and appropriate
conditions of pH and temperature. Depending on the starting substrates and the type of microbes,
thousands of different types of products can result. Generally, they fall into the categories of acids,
alcohols, gases, and a wide variety of organic compounds. Alcoholic fermentation of the type carried out by
yeasts yields primarily ethyl alcohol and carbon dioxide. A different fermentation pathway in Clostridium
botulinum yields complex proteins such as botulinum toxin.
First, it must be emphasized that there is no way to simply look at or smell a food product and be
able to tell if it contains toxins or other dangerous byproducts. They can only be detected by specialized
testing. Some food that smells or tastes disagreeable or unpleasant (for example, limburger cheese) may be
perfectly harmless and safe, and food that is contaminated or toxic often smells and tastes normal. The
bubbles of carbon dioxide coming off the fermenting wine are an expected product of fermentation and
harmless. Consequently, the inmates could not have known the wine contained something toxic until
someone drank it. In a way, they were serving as guinea pigs for their crude chemistry experiment.
The antitoxin is an antidote consisting of macromolecules called antibodies that bind specifically
to the toxin, an action which neutralizes or inactivates the toxin and prevents it from attaching to the
receptor on its target cells (the myoneural junction). If the toxin has already reached its target and bound,
the antitoxin is ineffective and the toxin will still have its full effect. So, the antitoxin must already be in
the bloodstream while the toxin is still circulating, which is within the first few hours of ingestion. After a
day or two, most toxin molecules will have bound to and entered the cells. There is also the effect of
quantity—it takes only a tiny dose of botulinum toxin to cause the disease, and the greater the amount
ingested the less effective the antitoxin will be.
The mistake made by the prisoners was that they added a chopped-up baked potato to the
fermentation substrate. And it was not a freshly cooked potato. Being baked, it still had its skin and had
been stored for several days in a tightly closed bottle. Clostridium botulinum lives naturally in soils all
around the world and could easily contaminate the surface of potatoes. It produces very resistant
endospores that can survive washing and even baking. It is also a strict anaerobe that germinates and grows
only in oxygen-free environments such as a closed bottle or fermentation container. It turns out that the
technique using yeast to make alcohol from the sugar in the potato was also an unfortunate recipe for
making botulinum toxin. Most pruno recipes use acidic fruits, which inhibit clostridial growth, and not
potato or other low acid vegetables.
Case Study 9 Perspectives
The source of the infection in the woman’s case was most likely the ventilator that controlled her
breathing. Medical devices are readily contaminated by patients, healthcare workers, and environmental
sources. When cases like this one are acquired during a stay in a hospital, they are termed nosocomial
infections. Such infections are most problematic in very compromised patients, although careful attention
to disinfection procedures and other precautions can greatly reduce their incidence. The second case
probably originated from the piece of shrapnel that picked up bacteria from the soil. Many other battlefield
cases were thought to have been acquired in field hospitals, but this was never completely clarified.
An opportunist or opportunistic pathogen is one that invades people whose defenses and
immunities have been compromised. For various reasons they have lost a protective barrier or lack an
effective immune response. Such patients usually have preexisting medical conditions such as cancer,
surgery, organic diseases, or traumatic injury. Because this category of patient is so susceptible, they can
become infected by common microbes such as A.baumannii from the environment, even those which are
not considered particularly virulent.
To say that a microbe has resistance to a drug means that it naturally possesses or has acquired a
genetic mechanism to avoid the effects of the drug. Usually, its genome carries one or more genes that can
eliminate the drug or prevent it from acting on the cells of the microbe. This leaves the microbe free to
infect and multiply even in the presence of that drug.
Most bacteria are rather efficient in transferring and acquiring genes from each other, and the
genes for drug resistance provide an adaptive advantage in an environment where these drugs are being
used. Such genetic recombinations take place in any habitat where microbes come in close proximity and
contact, including any colonized areas of the hospital and even the patient’s body.
Results from studies indicate that A. baumannii has extensive mechanisms for acquiring and
expressing drug resistance. It demonstrates all of the properties required to accept genes from other
bacteria, including pili for conjugation, DNA capture proteins involved in transformation, and
bacteriophages that act during transduction. This pathogen acts like a magnet for plasmids, transposons,
and integrons ( a large multidrug resistant plasmid) that encode multiple resistance genes. In one example,
A. baumannii had acquired an integron from a species of Pseudomonas carrying 45 genes that conferred
some type of resistance.
Once the pathogen has obtained these genes, actual contact with the drug will exert a selective
pressure on gene expression and favor the survival of a drug resistant strain. This may be repeated with
every drug used in therapy until the pathogen has extreme drug resistance. This is apparently what
happened with the patient in the first case--the bacteria had a hidden resistance gene that was selected for in
the course of the infection. Medical experts are alarmed that we may soon be facing a crisis with this
pathogen since there are few viable drugs and none in current development for this type of pathogen.
Case Study 10 Perspectives
Thousands of genetic disorders have been discovered in humans. When they are inherited from one or both
parents, they will occur in every cell of the body, because they are present in the egg and/or sperm. These
conditions, such as sickle cell anemia and cystic fibrosis, are termed congenital or inborn conditions and
are usually manifest early in life. Others (for example, some types of leukemia) are acquired through
mutations after birth and do not occur in all cells of the body. Some disorders are caused by a single gene
defect and others are caused by the malfunction of more than one gene. So far, geneticists have described
about 4000 single gene defects of the type in this case study. Most of them are rare in incidence, but
because there are so many different types, collectively they affect a significant portion of the population.
The DNA in a single human cell is an extremely long molecule—it would stretch 6 feet if the
DNA of all 46 chromosomes were unwound and laid out in a single line. Lying along this elongate
molecule are some 6.4 billion nucleotides that constitute the genetic code. Sequencing the precise order of
this code is somewhat simple in concept but rather difficult in practice. It usually requires cutting the DNA
in smaller fragments that can be analyzed separately. The order of a fragment is read one nucleotide at a
time by a method that can identify and differentiate each nucleotide by some unique characteristic. For
example, the Sanger sequencer uses a different fluorescent dye on each nucleotide that can be detected by
an electronic sensor that registers whether it is an A, G, C, or T. When numerous fragments have been
read, their codes can be used to reconstruct the codes of much longer stretches of DNA. Managing the
large amount of generated data requires computers that are capable of scanning the codes and determining
where particular fragments fit together and differentiating functional types of DNA, such as structural
genes, exons, and regulatory genes.
Recall that eukaryotic genes are split and not colinear; they cannot be translated directly into a
series of amino acids without processing. They are interrupted by short series of nucleotides called introns
that do not code for protein. The other intervening codes that do code for protein are called exons. Prior to
translation, the introns are excised by an enzyme complex, leaving only the exons for final translation into a
functioning protein. It is reasoned that, since a gene defect usually arises from a malfunctioning protein, it
will most likely arise from mutations in the exons and not introns. In addition, introns and other nonprotein-coding elements of the genetic code greatly outnumber the exons. As a result, concentrating on the
exons alone will prevent having to sequence billions of nucleotides that are probably not going to shed light
on abnormalities.
Bioinformatics is a relatively new science that uses computer programs to process and analyze
biological information, especially large complex sets of data that would be very difficult if not impossible
to manage any other way. It uses the tools of mathematics to organize and characterize important aspects
of biological systems as a way to provide deeper understanding of these phenomena.
Nicholas’s case is a good example of how it can be used. First, it was an essential part of the
genomics and sequencing, because only a computer could faithfully and rapidly store literally billions of
bits of data and thousands of pages of DNA codes. It was only through the completed human genome
informatics project that the doctors had a baseline with which to compare Nicholas’ DNA sequences. One
part of diagnosis is to use sequence alignment and comparison to track the exact position of genes and
exons. It also requires gene annotation, which guides the discovery of specific genes, polymorphisms, and
mutations of the type found in Nicholas’s case. Several specialized programs were developed to ultimately
find the single base substitution that could account for Nicholas’s genetic defect.
Case Study 11 Perspectives
Hepatitis C is the most common blood borne infection in the United States, with an estimated 3.2
million persons currently infected. This estimate is higher than the reported numbers of cases
because only about 10% of infected persons present with an acute case of the infection. These are
individuals who suddenly develop visible signs and symptoms indicative of a recent, diagnosable
infection. The remainder have a chronic infection, meaning that they have had it for a longer
period, generally lack symptoms, do not seek medical care, and are usually carriers of the
hepatitis C virus (HCV).
The three index cases identified in this case study had acute infections, implying that the
HCV had been recently acquired, most likely from a chronic carrier who had also been treated in
the clinic. What made it even more serious was that the circumstances of contact involved
thousands of patients, 90% of whom were at risk for developing a chronic infection. Finding and
alerting these patients was critical because they may develop aymptomatic infections and go on to
become carriers. Equally serious is the knowledge that people with chronic infections are also
prone to deadly liver diseases such as cirrhosis and cancer.
Most HCV infections of medical origin involve exposure to the body fluids or tissues of
another patient, although occasionally clinic workers are a source. Contact may occur when
errors are made during injections, biopsies, and other invasive procedures that increase the chance
of cross contamination. Most of the time, infection arises from contaminated equipment like
biopsy tools, endoscopes, needles and syringes, and injected liquids that are not properly
sterilized or handled. It is unlikely that one could get HCV from casual contact or environmental
sources.
Patients had to be screened for HBV and HIV because these viruses are also transmitted
by contact with blood. It is reasoned that people who are infected by one of these viruses have an
increased risk of harboring other viruses in their blood. Fortunately there were no cases of HBV
or HIV infections traced to the clinic among the 50,000 patients tested. Unfortunately, a single
patient became the source for 106 cases of hepatitis C, forcing the clinic into bankruptcy, and
costing the State of Nevada as much as 21 million dollars.
A detailed study ruled out all but one possible mode of transmission. It was traced to the
method used by an RN to draw up the anesthesia drugs from a vial into a syringe. He didn’t
understand one of the basic tenets of microbiology, which is that, just because you don’t see
evidence of contamination, there is always a possibility that microbes could be present, and that is
why one must always use scrupulous sterile techniques in any injections. Yes, the needle was
sterile, but the syringe was not. This is because when he injected a drug the first time, some of
the patient’s tissue fluid and blood was pulled into the space between the needle hub and plunger.
When this same syringe was used to give a second injection to the same patient, the common
practice of equalizing air pressure between the vial and syringe would have caused the mixing of
the patient’s blood with the drug in the vial. Then, when the vial was reused on a different
patient, he would have been injected with some of the viral-tainted blood. Even a tiny amount of
contamination is sufficient for this virus, which has an infectious dose of only 10-20 viruses!
Case Study 12 Perspectives
Health authorities estimate that between 600,000 and 800,000 needle sticks occur in medical
facilities per year. Exposure to HIV by this route has a potential of causing about 3 infections in 1000
people. The risk is high enough to warrant post exposure prophylaxis-- a preventive therapy to eliminate
the virus as early as possible. Prophylaxis can also be used pre-exposure to prevent possible infection
during a medical procedure such as dental work and some surgeries. If prophylactic treatment is
administered as soon as possible after post-HIV exposure, the drugs can act upon the virus before it gets a
chance to replicate inside host cells. Both of the drugs in combivir (retrovir and epivir) have the action of
preventing the HIV RNA from being converted to DNA and thus blocking further virus activity.
The idea behind combined therapy applies to both of the pathogens in this case. Both HIV and M.
tuberculosis have relatively high rates of drug resistance. Single drug therapy would select out a resistant
strain and allow it to flourish. But if two or more drugs are given together, any microbes resistant to one of
the drugs will be wiped out by another drug and visa versa. This effectively prevents the survival of any
microbes that are drug resistant. Often the drugs are administered in a 2, 3, or 4 drug “cocktail” that can
serve for either prophylaxis or treatment of infections. For HIV infection or AIDS, these drugs are selected
from one of the classes of reverse transcriptase inhibitors, protease inhibitors, and others (described in
tables 12.6 and 12.7). For tuberculosis, the choices are narrowed down to isoniazid, rifampin, ethambutol,
pyrazinamide, thiacetazone, para-amino salicylic acid, and streptomycin. Drug therapy for TB must also
include at least two of these drugs. Because there are emerging strains of TB that are MDR (multiply drug
resistant) and XDR (extremely drug resistant), it is necessary to do a culture and sensitivity test to ensure
that the selected drugs are actually effective.
This case was complicated by the AIDS patient having undiagnosed tuberculosis. His
pneumocystis pneumonia would have masked out the symptoms of TB and interfered with analyzing a
chest X-ray. Tuberculosis is notoriously slow to culture, and a few days or weeks are not enough time to
detect identifiable growth. Another contributing factor is that cutaneous tuberculosis is a relatively rare
infection and would not ordinarily have been suspected even from a skin puncture.
This case provides insight into several aspects of antimicrobial therapy. It shows that each
infectious disease must be treated on an individual basis with the drugs that have a specific mode of action
for the pathogen. Testing for sensitivity is especially important in selecting the best drugs and in detecting
instances of drug resistance. We have also touched on the issue of adverse side effects of drugs, which can
prevent compliance with therapy that is a major factor in the failure of HIV and TB treatment.
Case Study 13 Perspectives
The time from the first contact with the pathogen to the onset of symptoms is called the incubation period.
Salmonellosis shows a range of 36 hours because of variations in the infectious dose (number of cells
swallowed) and the medical condition of the host. The earliest symptoms to present during the prodromal
phase include nausea and vomiting; they are followed by a period of invasion marked by symptoms of
fever cases the manifestations subside within 1 to 2 weeks, which is followed by full-fledged cramping
diarrhea, and fever of the period of invasion. During the convalescent period, the manifestations subside,
but some patients continue to shed live bacteria even after the signs and symptoms are gone.
Evidence that differentiates sporadic from epidemic occurrence involves the number and location
of cases and whether they can be linked in any way. It became evident that this was an epidemic because
the number of cases rose by 10% of the usual number of cases in a relatively short period of time. This rise
was sustained for several weeks, and the cases came from the exact same source and serotype. The pattern
shown is consistent with a common source epidemic (figure 13.B). If the incidence had been sporadic,
there would have been a small number of isolated cases that involved different foods and unrelated
serotypes and would not have shown such a dramatic increase in incidence.
Transmission of salmonellosis frequently involves vectors. There were probably two different
types of vectors acting here. In one instance, bird or rat feces would both be likely reservoirs of
Salmonella. Because these animals harbor and nurture the microbe in their large intestine, they would be
considered biological vectors. The way that cockroaches passively transport the pathogen on the outside of
their bodies and spread it by contact puts them in the category of mechanical vectors. The investigators
were never able to make a final conclusion on the exact reservoir or which vector was more involved. It
was probably a combination of vectors and unsanitary production technology. They concluded that any
one of the violations seen by the inspectors could have caused gross contamination. The company did not
adequately separate raw and finished products and failed to take seriously the 12 times their product tested
positive for Salmonella by their own laboratory.
The infectious dose is relatively high because the microbe has to survive passage through the
stomach acid. Once they reach the small intestine, the pathogens adhere to the intestinal cells with their
flagella and are engulfed. Being gram negative, Salmonella species contain endotoxin in its cell wall,
which triggers inflammatory reactions that lead to the gastrointestinal disturbances and fever. In the more
severe cases, the infection becomes systemic as the pathogen enters the blood and perforates the bowel (see
figure 13.11d). In severely compromised patients, endotoxemia can lead to shock, circulatory collapse, and
death.
Case Study 14 Perspectives
Osteomyelitis is an inflammatory infection of the vascular spaces (marrow) within bones
(especially long bones and the ankle or wrist). It is frequently a focal infection that has arisen from a local
infection elsewhere in the body. It usually presents with an abscess forming in the tissue around the bone
and progresses to fever, chills, and pain at the site of infection. The swelling and inflammation of the
child’s bones were triggered by the Aspergillus hyphae growing into bone cavities. The presence of these
foreign cells initiated a series of local events, beginning with infiltration of phagocytes and other white
blood cells into the infected area. The white blood cells release biochemical mediators that induce classic
inflammatory signals of rubor, calor, tumor, and dolor. These mediators dilate blood vessels, creating
rubor or redness, which also has the effect of increasing the local temperature (calor). The reactions flood
the site with tissue fluid and cause swelling (tumor) and pain (dolor).
The major infectious agents that invade CGD patients are catalase-producing aerobic bacteria and
fungi. These types of organisms would usually be kept in check by normal phagocytes that literally douse
them in toxic oxygen intermediates such as superoxide ions, hydroxyl radicals, and hypochlorites. Because
CGD phagocytes lack these intermediates, they have little or no effect on organisms such as
Staphylococcus, Klebsiella, Aspergillus, and Candida. Viruses are not a significant cause of opportunistic
infections in these patients because viruses are not controlled by oxygen-based chemicals. Most other
defenses would be intact in CGD children, including innate barriers, the inflammatory response and
cytokines, interferon, the complement system, and T- and B-cell immunities, but they are not sufficient to
maintain full protection against some infectious agents.
Granulomas often develop at the sites of chronic inflammation. They consist of clusters of dead
tissue and immune cells surrounding a central core of macrophages that have been unable to control the
inflammation and have shut down (frustrated phagocytes). Even people with healthy phagocytes can
produce granulomas, but children with CGD produce them in pathological amounts because all of their
phagocytes are ineffective. The granulomatous nature of this disease is triggered by chronic inflammation
and build-up of dead neutrophils that could not completely finish the job of clearing the pathogen. When
the backup team of macrophages enter the scene to “clean up” the dead neutrophils and infection debris,
they are also unable to resolve the inflammation, and leave behind large inactive packets of cells that
remain in tissues for long periods of time. CGD patients suffer from recurrent opportunistic infections
because of this inability to clear the pathogen.
The immune stimulant gamma interferon has been beneficial in resolving infections in CGD
patients. This may be due to several factors. One contribution of interferon is to activate macrophages and
enhance the effectiveness of phagocytosis. Another is that it modulates T cells that can destroy infectious
agents without the need for phagocyte killing mechanisms. Interferon is often prescribed in combination
with antimicrobial drugs that target the pathogen. Since this disease is genetic, it must be treated
continuously with long term prophylactic medications and other therapies. There is no cure, but several
trials of gene therapy to correct the genetic defect are in progress.
Case Study 15 Perspectives
Rabies is a type of encephalitis in which the virus migrates by nerve cell processes from the portal
of entry, enters the brain, multiplies, and causes severe inflammation and damage to brain cells. Post
exposure prophylaxis is an immunization method designed to be administered as soon after contact as
possible. As we see in the two cases, its timing is critical. First the wound area is infused with human
rabies immunoblobulin (HRIG), a product purified from human serum, and HRIG is also injected into the
muscle of the arm. It contains high levels of specific antibodies that adhere to the virus and neutralize it.
This is a form of artificial passive immunity that will immediately deliver anti-rabies antibodies to the
wound and the circulation as well to keep the virus from invading the brain.
The rabies vaccine contains whole inactivated viruses grown in cell culture. The usual protocol
requires a series of four to five intramuscular injections that start on day 0 (first dose) and then proceed on
days 3, 7, 14, and sometimes 28. This form of immunization takes long but provides longer term artificial
active immunity that stimulates the patient to produce his own protective antibodies, T cells, and memory
cells. This will trigger a build-up in the number of antibodies in the blood and spinal fluid to eliminate the
virus.
The incubation period in rabies varies from two weeks to two months or more, depending on the
amount of virus and the distance it must travel to the brain. It was relatively short in both of these cases-around 2 to 3 weeks. The difference in outcome for both cases comes down to knowledge and timing. For
instance, the parents of the child in the first case did not know that the injury came from a bat, whereas in
the second case, there was no doubt how the child was exposed. This situation caused the delay in
treatment for child one, who had developed symptoms before it was started. In child two, it was started
before any symptoms appeared. Another significant factor was that child two had antiviral antibodies fairly
early and showed a rising antibody titer during a critical period. The antibodies must have developed a
high enough titer to control the virus. Keeping him sedated during this time and slowing brain activity may
also have been a factor in his survival. Child one already had such a high viral load in the brain that his
immune response could not catch up.
The main purpose of giving so many injections of vaccine is to flood the immune system with
antigen, which will expand the clones of lymphocytes that can react with this virus. This will result in
more activated B cells and plasma cells to make the specific antibody and more activated T cells to destroy
infected cells. Three doses are known to provide 100% protection, but additional doses are given to boost
the immune system. Every year in the United States about 40,000 people report potential exposure to a
rabid animal and are given PEP right away. The strategy has been so effective that not one healthy person
has ever gone on to develop a case of rabies after this treatment.
Case Study 16 Perspectives
A person’s ABO blood type is inherited as a combination of two alleles that code for glycoprotein
antigens on red blood cells. One allele comes from each parent, and the A or B alleles are both dominant
over recessive O alleles. Type A people inherit two A alleles or one A allele and an O allele; type B people
inherit two B alleles or one B allele and an O allele; type AB people inherit one A and one B allele, and
type O people inherit two O alleles. With the Rh factor, the Rh+ allele is dominant over the Rh- allele. An
Rh+ person inherits one or two Rh+ alleles, and an Rh- person inherits only Rh- alleles. Complications can
arise when blood is transfused because most people carry antibodies that react with blood antigens that are
different from theirs (an exception to this rule is type AB+). As long as a person does not receive blood of
the “wrong” antigenic type, these antibodies will not create a problem.
When an incompatible blood type is given, the donor RBCs are lysed by complement bound to
antibodies, which releases hemoglobin into the blood stream. The red hemoglobin molecules will change
the color of plasma and urine to a shade of pink. The disseminated intravascular coagulation is created by
RBCs being agglutinated into large clumps by IgM antibodies that clog up the small arteries and capillaries,
cutting off circulation to vital organs such as the heart, lungs, and kidney. The shutting down of his
circulation caused shock, low blood pressure, heart failure, and death.
It is generally thought that the plasma-borne antibodies against A and B antigens are produced
through a natural, normal process that unfortunately collides with medical science. There are glycoproteins
called heterophile antigens found in plants and bacteria that mimic the structure of human blood group
molecules. During early contact with these substances from food and normal microbiota, the immune
system is sensitized and makes antibodies that “lie in wait” in case of future contact with these antigens.
This is true only for the ABO system. Anti-Rh antibodies will only be formed if a person is exposed to the
Rh factor through a faulty transfusion or during gestation (see hemolytic disease of the newborn).
There are about 20,000 acute transfusion reactions per year in the US, with about 50 deaths. The
majority of these cases are attributable to staff errors in the hospital or bloodbank. A transfusion reaction is
a medical emergency. The treatment usually consists of flushing the blood stream with saline and
removing the hemoglobin from the bloodsteam with drugs. It is often necessary to give additional
transfusions to replace the entire contents of the bloodstream. The decision under these circumstances is
which blood to give safely when the actual blood type of the patient cannot be immediately clarified. This
is an example of a circumstance where type O- blood is considered for universal donation. It is safe to give
because it contains none of the A, B, or Rh antigens and will not trigger further hemolysis, agglutination, or
immune reactions.
Case Study 17 Perspectives
Bile is an agent added to media such as MacConkey agar that effectively selects for the growth of
enteric gram-negative rods. The rationale is that bacteria that frequent or occupy the large intestine are
likely to be resistant to bile since they are constantly in contact with it in this habitat. Bile is an effective
inhibitor of many species of gram-positive bacteria that might be found in fecal, food, and other types of
specimens. Another substance used to block the growth of gram-positive bacteria are dyes, which can be
used in differentiation as well.
The oxidase test detects the enzyme cytochrome oxidase, which is part of the electron transport
chain that adds oxygen to hydrogen ions and forms water. Because many bacteria lack this enzyme, it can
be helpful in the initial characterization of a genus. Fermentation tests detect the breakdown of a
carbohydrate in the absence of oxygen with the formation of acid, gas, and other products such as alcohols.
Taking into account the preliminary results, the most likely genera responsible for the outbreak were
Campylobacter and Vibrio, which share several traits. But the lack of spiral cells and having fermentative
and aerobic metabolism all narrow it to Vibrio as the causative genus.
The last series of biochemical tests confirmed that the pathogen isolated from patient specimens
and food was Vibrio parahaemolyticus. The growth requirement for salt, the fermentation of arabinose,
and being ONPG negative were key tests in this determination. Although all three Vibrio species live in the
ocean and are associated with eating raw or insufficiently cooked seafood, V. parahaemolyticus is the
leading cause of enteritis associated with shellfish consumption worldwide. Its relative V. cholerae is the
causative agent of cholera and V. vulnificus causes enteritis, wound infection, and septicemia.
The most specific evidence that verified the final identification were 1) serological tests using
slide agglutination (see 17.8b) and DNA fingerprinting of restriction fragments. The results showed all
isolates were serotype K6, which is the capsular antigen specific to V. parahaemolyticus.
It was essential to identify to species because the types of drugs and therapy can vary with the type
of Vibrio. It is also important to investigate the epidemiology of the infection so that public health officials
can trace back to the source and prevent further spread of the infectious agent. Investigators never
determined the exact source of the infection, since.V. parahaemolyticus was isolated from more than one
type of seafood. There was some concern that an asymptomatic carrier may have contaminated the food,
but no hospital workers tested positive for it. It appeared that shrimp or other seafood used to prepare
meals may not have been adequately refrigerated, allowing the pathogen to grow to infectious levels. This
was the first known instance of this pathogen being involved in a nosocomial setting.