Medical Significance of Pathogenicity Islands
Effects of horizontal gene transfer, seen from the viewpoint of diversity analysis of bacterial lineages, may seem a rather abstruse topic. Yet horizontal gene transfer among bacteria usually has immediate, practical effects on human health. Generally scientific studies have shown that not only is horizontal gene transfer spreading antibiotic-resistant genes, but horizontal gene transfer is also contributing to the evolution of disease-causing bacteria. A commonly seen recent example of a harmless bacteria devolving into a pathogenic one is the intestinal Escherichia coli O157H7 strain that occasionally causes fatalities. This “killer Escherichia coli” causes bloody diarrhea and kidney failure in children. Such strains appear to have been created by a horizontal gene transfer event from Shigella flexneri. The pathogenicity island, designated SHI-2 for Shigella, is a less virulent strain of E. coli that has acquired genes to enable it to make a new toxin. The toxin is thought to be responsible for kidney failure, a symptom not previously associated with E. coli infections.
As we develop a model of the origin of infectious disease from a creationist, biblical perspective, these bacteria may provide us with a glimpse of what has happened to living things over time in a fallen, cursed, and corrupted world. One helpful analogy is a faulty wire in a home. As any homeowner knows, failure to repair frayed electrical wiring is a fire hazard and can be disastrous. Any cut or a break in an electrical line can be very dangerous. One would suppose that the same holds true for a bacterium. Though electrical lines are safe most of the time, a corrupted line becomes a hazard.
Pathogenicity Islands and Other Prickles
In Genesis 3, the Scripture explains that when God cursed the earth, thorns and thistles appeared. These “prickles” came about because of man’s sin. From this point onward, we might imagine that genes were corrupted and germs were formed. In a figurative sense, what was previously “round” became sharp. The sharpness of bacteria (and other microbes) in today’s terms has to do with pathogenicity factors–namely, genes that code for poisons, toxins, enzymes, proteins, and virulence factors.
What makes a bacterium pathogenic? As mentioned before, bacteria in the body and soil were originally made for beneficial purposes. However, after the Edenic curse, beneficial structures deteriorated into malevolent structures through means of mutation, retrogression, and contamination. Smooth, round leaves and stems became thorns and thistles. Some of the contaminating processes that lead to pathogenicity include plasmid transfer, transduction, and conjugation.
While studying anthrax, Robert Koch was the first person to discover direct evidence that bacteria caused disease. Robert Koch first defined pathogenicity in 1890, when he extended his germ theory. These “pathogenicity islands” convey the virulence that is necessary for disease to develop. Since then, hundreds of pathogenic bacteria have been associated with disease. Pathogenicity islands have been used to refer to the clusters of genes responsible for virulence (e.g., enzymes and toxins), and they are fairly large segments of the genome of the pathogenic strains that are absent in nonpathogenic strains. These islands have many of the properties of intervening sequences, which suggests that they move by lateral gene transfer. These blocks of genetic information could move from a pathogenic strain into an avirulent organism, and then convert it to a pathogen. Such lateral transfer processes show how the corruption of a pure “kinds” lineage can make quick jumps and a break in the Creator’s original plan.
Pathogenic bacteria are defined by the capability of causing disease in a host. What makes a bacterium pathogenic is the set of genes that it possesses. The genes can be distributed on the chromosome, on plasmids (independent extrachomosomal genetic elements) and on a prophage. A prophage is a bacteriophage in which latent viral genes are incorporated into the bacterial chromosomes without causing disruption of the bacterial cell. Recently, blocks of virulence genes in the chromosome have been observed in virulent strains but not in avirulent strains. For example, normal E. coli does not possess these blocks whereas pathogenic E. coli does. These significant differences (e.g., nucleotide composition) between the chromosome and the pathogenicity island are observed and typical; it suggests that mobile genetic elements may have played a significant role in the devolution of pathogens.
One line of evidence for horizontal gene transfer as a selection force comes from examination of bacterial genome sequences. To find genes with two very similar DNA sequences in two distantly related bacteria suggests that these sequences might have been acquired by horizontal gene transfer. The number of such examples is increasing. The growing appreciation of how much horizontal gene transfer may have contributed to the variation (i.e., microevolution) of bacteria has caused microbiologists to question whether the “standard evolutionary tree,” which is characterized by well-isolated branches, should in fact be replaced by interwoven nets of branches. If two species have been differentiated on the basis of differences in their ability to utilize lactose and whether the genes that allow lactose utilization can be transferred by horizontal gene transfer, how firm then is this definition of species? Is this not variation with a “kind” or type? The significance of horizontal gene transfer is that it represents a defilement of the original bacteria types.
Mobile Genes Pose a Problem of Defilement
The concept of keeping the “seeds” separate and distinct is very insightful on a variety of microbial genetic issues. In Deuteronomy 22:9 and Leviticus 19:19, God tells the Israelites that if they do not keep the cattle separate from “diverse” kinds and seeds separate from “diverse” seeds, they will receive a curse on not only their immediate field, but also their vineyard. You cannot help but wonder if God meant that He knew the natural consequence of diverse gene mixing that leads to disease. It is clear that mixing genes from separate “kinds” (or types) is not the Creator’s way.
One application of this principle might render the passage as “Do not randomly plant two kinds (strands) of genes (DNA/RNA) in your cells; if you do, not only the microbes you ‘plant’ in the nucleus and plasmid but also the fruit of the entire host (man and animals) will be defiled (with virulence).” The exception to the consequence of diverse gene mixing is genetic engineering, because that is designed and controlled gene mixing. Defilement or corruption can be seen in pathogenicity islands that are added (via horizontal transfer) from one bacteria kind to another. This principle of randomized gene mixing can be seen in the transfer of pathogenicity islands in bacteria and in the origin of avian flu through the mixing of bird and mammal influenza strains.
The Origin of Bird Flu
Bird flu poses one of the greatest threats to human health if a pandemic occurs. Recall from Chapter 8, the Influenza A virus is an RNA virus that can change quickly. This rapid change occurs through the accumulation of small point mutations called antigenic drift, as well as through major genome reassortment called antigenic shift. Viruses rapidly change primarily because of the presence of many error-prone viral enzymes that replicate their genomes. This is especially the case with RNA viruses. Consequently, RNA viruses have mutation frequencies that are several folds higher than the observed mutation frequencies of DNA viruses.
Animal viruses may be degenerate genes coated by proteins that are cobbled together by co-option processes, or they may be decaying products from several genomes and proteins models. Recall from Chapter 7 the specific example of the degeneration of a normal gene (i.e., retrotransposons) to a retrovirus. On human chromosome 22, a gene (i.e., human endogenous retrotransposon) turns off the immune system during pregnancy. This design prohibits the mother’s immune system from damaging the child’s body. These genes cannot fully replicate, only infecting local immune cells (such as macrophages) of the uterus, thereby preventing them from initiating a full-blown immune response. Thus, the mother’s immune system remains able to respond to other infections but is specifically prevented from raising an immune response to the developing embryo. So in creation, the selective ability to turn off the immune system for protection would be a “good” design. However, since the corruption of creation, the degeneration and corruption of good genes in human and animal hosts have changed.
Perhaps these genes then co-opt, or randomly acquire proteins, and then leave the body as pathogenic viruses. These pathogenic viruses then open the world to devastating infections.
Influenza A Viruses
One might imagine that at one time good genes degraded in wild birds (i.e., ducks), but it did not affect them. However, once these deteriorated genes left the birds and transferred to mammals (e.g., pigs) and man, the major problems emerged. The terrible Spanish flu (1918–1919) developed quickly and mysteriously. It probably picked up some key genes from an avian source (possibly a duck or goose). Analysis by Dr. Jeffery Taubenberger and colleagues indicates that the H2N1 flu virus originated in an avian flu strain but spent time “evolving” in an unidentified host before emerging in 1918. It appeared mysteriously out of nowhere, caused tremendous distribution, and went away just as mysteriously when World War I ended. This was a corruption and decay of the originally good design of gene transfer, re-assortment, and recombination.
All known influenza A strains that infect humans are perpetuated in aquatic birds where it appears to cause asymptomatic infections. From these aquatic birds, influenza virus particles are excreted in high concentration in their feces, which helps spread the virus between birds and mammals. Besides residing within wild aquatic bird reservoirs such as ducks, two groups of influenza A viruses (the H5 and other strains) can infect domestic fowl. These strains are responsible for causing avian flu, which lately has received a lot of media attention because of its presence in chickens that are raised for human consumption. As well as infecting birds, two subtypes of influenza A can also be isolated from swine, which may be referred to as swine flu subtypes. These two strains isolated from swine are the H2N1 and H3N2 subtypes, with the former being a human pandemic strain.
Influenza in Birds Causes Little Harm
Influenza A viruses are brought about in the wild birds of the world, predominantly in waterfowl, in which the 16 subtypes coexist in perfect harmony with their hosts. In these natural hosts, the viruses remain stable in molecular form, showing minimal change (variation) at the amino acid level over extended periods. This fact indicates that the influenza-bird association is ancient; this lack of change is surprising because influenza viruses are segmented, negative-stranded RNA viruses that have no quality-control mechanisms during replication and are highly prone to variation. After transfer to a new type of host, either avian or mammalian, influenza viruses undergo rapid corruption, devolution, or defilement. However, all 16 HA subtypes, including H5 and H7, have, until recently, been considered to be benign in their natural hosts. This benign equilibrium between the influenza virus and its host may have changed.
The Genesis of Bird Flu
Before 1997, no evidence had indicated that H5 influenza viruses could infect humans and cause fatal disease. The precursor of the H5N1 influenza virus that spread to humans in 1997 was first detected in China, in 1996, when it caused a moderate number of deaths in geese. However, it attracted very little attention at the time. This goose virus acquired internal gene segments from influenza viruses later found in quail. It also acquired the neuraminidase gene segment from a duck virus. This all took place before the goose virus became widespread in live poultry markets in Hong Kong and killed 6 of 18 infected persons. Culling all domestic poultry in Hong Kong destroyed this H5N1 virus; the genotype has not been detected since that time. However, different reassortments continued to emerge from goose and duck reservoirs that contained the same H5 HA glycoprotein but had various internal genes. The H5N1 viruses continued to change, and in late 2002, a single genotype was responsible for killing most waterfowl in Hong Kong nature parks. This genotype of H5N1 spread to humans in Hong Kong in 2002, killing 1 of 2 infected persons, and was the precursor to the current strain of bird flu (i.e., the Z genotype) which has become dominant. The current strain spread in an unprecedented, first-time fashion across Southeast Asia and is now spreading across Europe and parts of Africa.
To date, more than 140 million domesticated birds have been killed by the virus or culled to stem its spread; as of spring 2006 more than 150 persons have been infected in Asia, Africa, and Europe. These recent H5N1 influenza viruses have acquired the unprecedented and disturbing capability to infect humans. These incremental changes intensify concern about this bird flu virus’ potential to cause a global pandemic. These genes were presumably acquired from viruses found in waterfowl in Southeast Asia, but the actual gene donors have not yet been identified. While most H5N1 influenza viruses that have been isolated from avian species in Asia since 1997 are highly pathogenic in certain poultry (pheasants and domestic fowls), they show varied pathogenicity in other species.
There is evidence that swine also may be involved in the interspecies transmission by providing an environment where human influenza viruses originate. Pandemic subtypes rarely occur, yet when they do emerge it happens suddenly. Many of these subtypes first appeared in China. It is believed that this is because China’s cultural heritage allows considerable domestic exposure to aquatic fowl, such as ducks, as well as a close domestic association with pigs. Influenza viruses can use a process known as genetic reassortment to create new pandemic strains that represent a combination of avian, human, and possibly even swine genomes. In reassortment, parts of individual genomes between closely related strains of virus are exchanged, creating a viral strain that is capable of infecting a different host, such as humans.
Making the Jump from Chickens to Humans
Currently, there are three potential pandemic influenza A strains (H5N1, H7N7, and H9N2) with all three being recently isolated from domestic fowl. The H9N2 strain first emerged in Hong Kong in 1999 and recently reemerged in 2003. This particular strain has killed three people and caused the mass killing of chickens in Hong Kong poultry markets to rid the influenza. Similarly, the H7N7 strain emerged in the poultry industry in the Netherlands. The H7N7 strain of influenza A has killed one person, and this has also required the slaughter of chickens by the thousands. Ominously, evidence was obtained for H7N7 human-to-human transmission as well as infection within nearby pig farms. In 2003, the H5N1 subtype emerged in Asia, and has now proven fatal to people on three continents and caused the culling of over 200 million birds.
As of spring 2006, not one of these viruses appeared to have acquired the necessary characteristics to cause widespread human infections, although experts seem to agree that it is only a matter of time before this happens. Of the three subtypes, perhaps the H5N1 subtype has the greatest pandemic potential. A hyper-virulent-H5N1 strain now is endemic to Asia, and unfortunately, China has recently reported that the highly virulent H5N1 avian flu strain can now be isolated from pigs. These observations make it likely that reassortment with a human strain will occur to produce an H5N1 virus with the capability for widespread human-to-human transmission. Therefore, the United States government and other international agencies have declared the development of a vaccine to protect against the H5NI strain a top priority.
The “Spanish” influenza pandemic remains a threatening warning to public health. Many questions about its origins, its unusual epidemiologic features, and the basis of its pathogenicity remain unanswered. The public health implications of the pandemic therefore remain in doubt even as we now grapple with the feared emergence of a pandemic caused by H5N1 or another virus. However, new information about the 1918 virus is emerging, for example, sequencing of the entire genome from archival autopsy tissues (see Historical Focus 9.5). But the viral genome alone is unlikely to provide answers to some critical questions. What caused so many deaths and severe infections? Historical Focus 9.5 discusses the 1918 flu pandemic and attempts to provide an understanding for this specific influenza. It also attempts to provide implications for future pandemics that require careful experimentation and in-depth historical analysis.
Mechanisms of Spread
Were the highly pathogenic H5N1 viruses transferred within and between countries by persons, poultry, or objects that carry germs (i.e., fomites)? In previous outbreaks of highly pathogenic H5 and H7 infection in multiple countries, the spread was directly attributable to humans. The primary means in which influenza virus is spread in poultry is by the movement of poultry and poultry products; establishing good security measures on poultry farms is therefore an important defense. The poultry industry is a huge, integrated complex in Asia. Nonetheless, the supposed involvement of multiple lineages of H5N1 argues against human-mediated spread from a single source. Live poultry markets are an amplifier and reservoir of infection, and they probably play a role in the maintenance and spread of the virus in the region.
However, a number of other factors unique to affected Asian countries make control difficult. Backyard flocks are common in the region, and these domesticated birds are not subject to any security measures. Fighting cocks are prized possessions and are often transported long distances. Fighting cocks may also play a role in the spread of infection and in transmission to humans. Many of the affected countries have weak veterinary communications and are facing highly pathogenic avian influenza outbreaks for the first time. The migrant ducks that commonly wander through rice fields scavenging fallen rice seeds are another potent mechanism for the spread of infection.
Keep the Seeds Separate and Distinct
“Thou shalt not let thy cattle gender with a diverse kind: thou shalt not sow thy field with mingled seed” (Lev. 19:19, KJV). One modern paraphrase of Leviticus 19:19 might be “Don’t mix your domestic livestock (i.e., pigs) with wild birds (i.e., ducks) or they may become virulent.” Later, the Bible says “Thou shalt not sow thy vineyard with divers seeds: lest the fruit of thy seed which thou hast sown, and the fruit of thy vineyard, be defiled” (Deut. 22:9, KJV). This principle of defilement or corruption may explain the origin of avian flu through the mixing of avian and mammal influenza strains. Farmers in areas where bird flu is endemic should be careful about allowing their domestic livestock to intermingle with wild birds. In addition, they should keep a clear separation between human and animal inhabitation.
Historical Focus 9.5
Doughboys Die of the Spanish Flu
America’s soldiers of World War I were referred to as doughboys. In 1918, many army privates were destined for the trenches of World War I in France. Some otherwise healthy young men began to bear symptoms of the flu. The Spanish flu was unusual because the most susceptible people were young, strong, and in the prime of life. The 1918 influenza victims were from healthy 20-to-50-year-old age groups (i.e., the age of military service), as opposed to the normally more susceptible age groups of the very young or very old. Uniquely, it produced a deep cyanosis (blue skin) that affected the face, lips, and lungs. Somehow, the virus penetrated the deepest parts of the lung for unknown reasons. It was known to kill some people in as little as 12 hours after contracting the virus. There was also an extraordinarily high mortality rate of 20 times the norm for influenza. In many of the cases that did survive the critical first few days of the influenza attack, death was precipitated by a rampant secondary infection with pneumonia bacteria.
In particular, one 21-year-old complained of chills, fever, headache, backache, and a cough. Within a week, he was dead—1 of 21 million people worldwide who would succumb to the influenza pandemic of 1918. For almost 90 years, samples of the doughboy’s lungs sat in a warehouse run by the Armed Forces Institute of Pathology in Washington, D.C. RNA bearing the solution to an enduring mystery lay hidden in his tissues; this RNA contained the genetic code for the worst pandemic in human history. Dr. Jeffery K. Taubenberger and his associates exhumed the deadly flu strain and fragments of its genes from the frozen tissue of the buried doughboy. Taubenberger and his colleagues had begun their search for the virus’s genes by selecting at random 28 of the 70 pandemic victims whose lung samples are stored at a government facility. Autopsy reports from 1918 disclosed that seven of these servicemen died soon after becoming ill, which enhances the likelihood that lung tissue might contain intact bits of RNA from the virus’s unusual eight-strand genome.
People who live longer are less likely to harbor the virus, because the body’s defenses eradicate the microbes, Taubenberger says. In such cases, bacterial pneumonia delivers the fatal blow. But in the seven servicemen who died quickly, the immune counterattack might not have had time to wipe out the virus. The researchers drew a blank in six cases. This Army private, however, was unusual. His left lung had suffered extensive bacterial pneumonia, but his right lung had not. (The most common cause of actual death from influenza was not the virus itself but rather by pneumonia. The most common form of bacterial pneumonia is caused by the bacterium, Haemophilus influenzae.) This raised the possibility that the right lung might still harbor the virus. To find out, the researchers removed some tissue from the paraffin in which it was stored. Step by step, they broke it down until only RNA remained. It yielded valuable information and could be helpful if a future pandemic occurs. Dr. Taubenberger believes that it could happen again.
Virologist Robert Webster of St. Jude Children’s Research Hospital in Memphis, TN, thinks that the 1918 virus represents the ultimate disease-causing agent. We need to understand as much as possible about this virus because the world will get another pandemic, maybe late in this century or early in the next. One pandemic was one too many. In the 1918 outbreak, nearly 700,000 people died in the United States alone. Over 50 million worldwide died of the Spanish flu over a two-year period. This was the pandemic of the century!
The people who preserved this tissue probably never imagined what might be possible down the road. Dr. Ann H. Reid, who worked with Dr. Taubenberger, made millions of copies of nine RNA fragments of five flu genes. Dr. Thomas G. Fanning then deciphered the sequences of the fragments and compared them to every other known sequence of the flu gene. It is unique and was designated H2N1. The team has also confirmed prior evidence suggesting that the sequences most closely resemble those from swine flu. Researchers are attempting to rebuild the entire genome of the virus, perhaps yielding clues to its hypervirulence. Many researchers feel that its information might help in the making of a vaccine, if needed. According to Dr. Webster, if this virus strain were to reemerge, this information would serve to get a best-match vaccine that would probably protect us quite well. In conclusion, the understanding of the 1918 virus (arguably the worst pandemic of all time) may help us to deal better with epidemics of the future, including avian flu and other virally caused diseases, like SARS. The scary thing about the 1918 influenza epidemic is that it came without warning, then affected young men who were uncharacteristic victims of any flu known and mysteriously went away. Some worry that avian flu may do the same.
Future Flu Pandemics
Over the years, several pandemic viral strains of influenza A have arisen. For example, there was the 1890 flu, the 1918 Spanish flu, and other flu epidemics in 1957 and 1968. Therefore, it appears almost inevitable that the next pandemic strain is currently lurking in some reservoir. Despite considerable knowledge of the genetic structure of the 1918 virus, no one is quite sure why the flu that year was so virulent, and there is still considerable scientific disagreement about where it came from. It is likely that a new surface protein appeared on this virus, one that humans had never encountered before and to which they therefore had no immunity. But there must have been other genetic features of this virus that also contributed, and these remain a mystery, even though complete viral gene sequences of the bug have been developed from the preserved tissue.
So the question arises: Could it happen again? In a sense, it already has happened again, though not in such a devastating form. The “Asian flu” of the winter of 1957–1958 killed 70,000 Americans. The 1968–1969 “Hong Kong flu” killed 34,000. The new subtype that appeared that year, called Influenza A (H3N2), was milder, perhaps because only the hemagglutinin protein changed and the neuraminidase stayed the same, therefore people may have had some residual resistance to it. Still, Influenza A (H3N2) has caused 400,000 deaths in the United States since its emergence in 1968, and 90 percent of them were among people older than 65. In fact, the subtype that caused the 1918 pandemic, Influenza A(H2N1), continued to circulate for years, having undergone antigenic drift. This caused a large number of deaths in 1920 (probably reduced by greater immunity among the population as well as by the attenuated form of the virus) and then periodic outbreaks until it disappeared in 1957. None of the outbreaks, though, was anywhere near the severity of that of 1918. What made the 1918 strain so deadly has been a longstanding medical mystery. Analysis for those genes and proteins revealed viral features that could have both suppressed immune defenses and provoked violent immune reactions in victims, which contributed to the high mortality. Known bird and mammal influenza hosts are unlikely sources of the pandemic virus, so its origin remains unsolved. In 1977 it reappeared, but again it was without devastating consequences.
In any case, most experts on the flu think that the answer is yes, it could happen again. The reservoir of influenza is believed to be wild waterfowl, and the pandemics of 1957 and 1968 were both produced by changes in the surface proteins of an avian strain of the virus. Such an alteration in the future could produce a pandemic as bad as, or worse than, the 1918 disaster. Of course, circumstances now are a bit different. We have a flu reporting system; we know how to make vaccines against the flu; we have antibiotics to treat the bacterial infections that killed so many in the pandemic of 1918; we even have several antiviral medicines that are helpful. But we also know that microbes are persistent and very adaptable, and that there is always the possibility that one of them will come up with an answer to the most elaborate defenses that have been constructed against it.
The Future of Bird Flu
At the time of this writing, bird flu appears to be imminent to spread into the United States via the Alaskan Aleutian islands. As birds fly naturally across the Bering Strait from Siberia, some birds will inevitably carry with them avian flu. Once it lands in North America, the fear is that it will spread down the west coast. The US government is monitoring it closely. However, the predictability is like forecasting a hurricane when a tropical storm is in the Atlantic Ocean. You know that it is coming, but whether it will be a Category 1, 2, 3, 4, or 5 is known only to God before it actually hits. The heightened monitoring of avian flu will help warn us of a potential epidemic. But what can we do? Perhaps, regulations on keeping livestock separate from wild animals may reduce “viral” mixing. In addition, we can stockpile antiviral medications and vaccines. Beyond this, it is in the Creator’s hand. Concerns about future outbreaks of avian flu (mutation) and other influenza pandemics will continue. More research needs to be done on the origin of infectious disease from a creation, biblical perspective. There are still many mysteries and challenges ahead.