Entomopathogenic nematodes and Their Enterobacteria Symbionts as agents of Termite Control

— This article by Jerry Cates, first published in August 2001, was last revised on 07 December 2013. © Bugsinthenews Vol. 02:08.

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This article discusses the biology, anatomy, and behavior of entomopathogenic nematodes (EPN) and their bacterial symbionts, specifically as agents of subterranean termite control. Particular attention is paid to the utility of EPN for termite colony interdiction when applied in conjunction with nematode-optimized termite interceptors.

Introduction:

Nematodes are roundworms, or threadworms, in the phylum Nematoda. Some species live as parasites inside the bodies of insects and other organisms, often with no observable effect on their hosts. Others are known to produce a broad range of noticeable sequelae, ranging from minor discomfort to disease and death.

Entomophilic nematodes, i.e., those which have affinities for insect hosts, have been the subject of scientific inquiry since 1602, when Ulisse Aldrovandi, the Italian botanist who created the botanical gardens of Bologna, found and documented the emergence of worms from the carcasses of dead grasshoppers. Among the long list of entomophilic nematodes are found a somewhat smaller grouping of entomopathogenic nematodes that, in addition to having a natural affinity for insects, also produce pathological conditions such as those observed by Aldrovandi. Lacey et al. (2001) lists as many as 30 families of nematodes that are associated with insects. Scientists have focused on seven of these for their insect associations, and on other nematode families for their pathological associations with non-insect pests.

For example a nematode in the family Rhabditidae, Phasmarhabditis hermaphrodita, infects slugs in the genus Deroceras. Kaya (2000) reported that P. hermaphrodita controls many slug and snail species with an efficiency equivalent to chemical control standards but without the adverse effects on non-target mollusks characteristic of chemical-based controls. Further, nematodes from two other families also show important molluscicidal activity against two slugs of economic importance, Deroceras reticulatum and Limax marginatus.

Many successful uses of EPNs to control insect pests have been demonstrated. Deladenus siricidicola, an EPN in the family Phaenopsitylenchidae, successfully controls the woodwasp Sirex noctilio. When inoculated along with Oryctes nonoccluded virus, and Entomophaga maimaiga, a fungus, this same EPN successfully provides long-term suppression of the palm rhinoceros beetle Oryctes rhinoceros, and the gypsy moth Lymantria dispar.

Romanomermis culicivorax Ross & Smith, an EPN in the family Mermithidae, successfully suppresses mosquito larvae and recycles at high levels in suitable habitats. Only a low tolerance of conditions prevailing in certain host habitats, and the fact that Bacillus thuringiensis israelensis (Bti) provides equivalent control at less cost, prevent this EPN from being used extensively for mosquito control.

Of the six nematode families remaining, three have complicated life cycles that make mass production difficult. These families, the Tetradonematidae, Allantonematidae, and Sphaerulariidae, are undergoing further study in a number of scientific laboratories and may prove useful in the future.

The three remaining nematode families have demonstrated great economic success in insect control programs, due to a combination of host specificity and virulence, combined with ease of mass production. EPN from two of these families cause host death soon after entering the insect’s body (the Steinernematidae and Heterorhabditiae), while those from the third kills its host later, upon emerging from the host’s body (the Mermithidae).

Regulation of Entomopathogenic Nematodes:

Entomopathogenic nematodes are exempt from regulation by the U.S. Environmental Protection Agency. These organisms are known to kill insects, including termites, and have been used to fight agricultural pests for decades. As such, nematodes provide one of the most important methods of pest management on farms in America and throughout the world. Their role in reducing the use of agricultural chemicals, and lowering exposures to hazardous pesticides for farm workers and consumers, cannot be overstated.

Nematodes used in insect control produce mortality only in insects, though they are known to produce morbidity in certain other invertebrates. Though otherwise harmless to mammals, at least one strain of EPN that is not used for insect control has been found, under extremely unusual conditions, to produce morbidity in mammals. This nematode, a close relative to Heterorhabditis indica, was identified in Australia as the carrier of a mammalian pathogen, Photorhabdus asymbiotica, var. Kingscliff, that apparently caused an isolated number of cases of human morbidity there and in the United States.

Though the discovery of Photorhabdus asymbiotica, var. Kingscliff does not tarnish the stellar safety record of EPN used as biological agents for pest management today, it illustrates an essential truth that should always be kept in mind: a measure of caution is always wise, even when dealing with materials that are vouched for as safe by the most trusted of authorities.

Using EPN for Termite Control

Certain entomopathogenic nematodes are virulent enough to produce 100% mortality within an inoculated termite colony. As with chemical control methods, however, their virulence is limited to the point that a single inoculation of large termite colonies, at only one inoculation point, is generally insufficient to produce absolute colony elimination. Nullification of the typical termite colony, to the point where it is no longer able to infest and damage manufactured structures and botanicals such as living trees and shrubs, can only be assured through multiple EPN inoculations of a plurality of termite infestations within the termite colony’s foraging zone,  carried out over a period of several months.

Termite colony elimination using EPN is presumed to have occurred when, after conducting a series of EPN inoculations as described above, all termite foraging activities within the monitored zone have ceased for at least twelve months. Afterward, the areas within the zone at risk for future termite infestation, must be monitored on a regular basis, to detect the presence of new termite colonies, arriving from the unmonitored periphery, so that they, too, may be inoculated with EPN. In addition, all at-risk cellulosic matter within the monitored zone may be treated with a termiticide capable of producing mortality in termites that arrive in the future and feed on the treated materials.

EPN exhibit a delayed mode of action

Delayed morbidity and mortality are important features of nematode infections. Termite workers do not stop moving except to feed on raw cellulose and, on occasion, to rest; when at less than optimum health, they “walk it off” and feed on recent fecal deposits, in the colony’s workings, left by their cohorts, rather than stand idle. This adaptation ensures their intestinal tract is replenished with a constant source of fresh gut fauna, and that any food-related toxicant is passed out of the gut quickly.

While effectively curing many if not most natural intoxications from contaminated food consumed by worker termites, the adaptation described above is ineffective at resolving an infection from an EPN, and in fact assists in propagating the EPN infection within the termite colony. Once a termite is so infected, it continues to travel within the termite colony’s workings for hours to days (depending on ambient temperatures) afterward, potentially distributing nematodes far beyond the inoculation site. When the infected termite expires, its cadaver does not putrefy, and thus does not produce any of the chemical markers that are used by uninfected nest-mates as a signal for them to remove the corpse from the colony’s workings.

Termite workers that encounter a putrefying corpse within the colony workings immediately transport it to a point outside, either by cutting a hole in a weak point in the workings, and pushing their dead nestmate out, or by placing the corpse in a dead end tunnel and sealing it off from the rest of the colony. That instinctive behavior is a response to the secretion, by the corpse, of specific products of putrefaction. For reasons described below, termites killed by EPN do not putrefy, so their living nestmates are not signaled to discard their bodies. Instead, the cadaver remains inside the colony’s functional superorganism, where it incubates a new batch of nematodes. These later emerge from their cadaverous incubation chamber, to ambush and infect other members of the termite colony.

Though initial exposure to the inoculum in the termite interceptor involves mostly workers, all castes in the termite colony are susceptible to nematode infection. As cadavers in the workings of the colony’s superorganism release new batches of infectious juveniles (IJ’s), any termites passing nearby — including reproductives and soldiers — become targets for new infections.

Nematodes do not appear to elicit complicated, instinctive, avoidance responses in individual members of termite colonies. Studies have shown that many termite species, such as Heterotermes aureus, are unable to detect nematodes and take no evasive actions in their presence. Other studies show that at least three termite species, Reticulitermes tibialis, R. speratus, and Coptotermes formosanus, detect the presence of nematodes in their colony’s workings and take simple, though ineffective, evasive action as a result.

A study by Wu, et al., observed that, even though evasive action was taken by members of C. formosanus and R. speratus colonies, the response was so ineffective that nematode infections occurred anyway. Though the total number of EPN infections are fewer with those termite species that exhibit complex avoidance responses, repeated inoculations of nematodes into their colonies, via specialized EPN-optimized termite interceptors and inoculators, have been shown to be capable, over time, of overwhelming and eliminating their termite colonies.

Phoretic Bacteria and Entomopathogenic Nematodes:

Bacteria of the genera Xenorhabdus and Photorhabdus live in a phoretic relationship with certain nematodes and as pathogens in the insects the nematodes invade. The definition of the bacteria-nematode relationship is both phoretic (with respect to the bacteria, which use the nematode as its vehicle to gain access to an insect’s blood supply, or hemocoel) and symbiotic (with respect to both the nematode and the bacteria, each of whom benefits from their close relationship). Nematodes need the bacteria to survive, as they feed on the bacterial mass that the infection produces. The insects invaded by the nematodes soon die from infections caused by the bacteria the nematodes bring with them.

Species of these bacteria are rod-shaped, facultative, anaerobic, Gram-negative members of the bacteria family Enterobacteriaceae. Members of the Enterobacteriaceae that serve as symbionts in entomopathogenic nematodes (EPN) are normally harmless to humans and their pets. Medical records from the American Civil War and World War I attested to the fat that when a certain species (now known as Photorhabdus luminescens) contaminated human wounds, the wounds–which glowed faintly in the dark — often healed faster than normal wounds not contaminated with the bacterium, presumably because the P. luminescens bacteria secreted broad-spectrum antibiotics that inhibited the development of harmful microbes.

Though similar to other Enterobacteriaceae, species of Xenorhabdus and Photorhabdus tend to be bigger and, unlike most other members of this family, do not reduce nitrates to nitrites. Some emit light during portions of their life cycles, a feature that makes them unique in a number of ways. Only three genera of bacteria are known to luminesce. Two are found in sea water and marine organisms, but the terrestrial bacteria with this property are all members of the genus Photorhabdus (recently reclassified from a single species, Xenorhabdus luminescens, to this new genus, originally believed to contain a single species but now recognized to comprise several.)

Microbiologists studying the DNA of various strains from these genera are beginning to find evidence that they have been around for a long time. For example, after analyzing 76 different Xenorhabdus strains from 27 species of Steinernema nematodes collected in 32 countries, one investigator (Tailliez 2006) found strong evidence that the common ancestor of the genus emerged between 250-500 million years ago.

Xenorhabdus and Photorhabdus species kill insect hosts so quickly that nematodes carrying them don’t have to adapt to the insect’s life cycle. That makes their EPN involvement effective against a large number of insects, including, under certain circumstances, subterranean termites.

Nematode/Bacterium Specificity:

The specificity of the bacteria/nematode association appears to be an important safety feature regarding all EPN. As a result of this specificity, the risk that any nematode species whose symbiotic bacteria does not, today, produce morbidity in humans, will become a human pathogen in the future, appears to be comfortably remote.

The safety of EPN with respect to mammals and non-target organisms is well-documented. Ralf-Udo Ehlers, a recognized authority on biological control of insects, describes in a paper written in 2003 the history of the use of these biological control agents:

“Since the first use of the entomopathogenic nematode Steinernema glaseri against the white grub Popilla japonica in New Jersey (USA) (Glaser and Farrell, 1935), not even inferior damages or hazards caused by the use of EPN to the environment have been recorded. The use of EPN is safe for the user. EPN and their associated bacteria cause no detrimental effect to mammals or plants (Poinar et al., 1982; Boemare et al., 1996; Bathon, 1996; Akhurst and Smith, 2002).”

None of the EPN presently in use as agents for the control of insects, or their associated phoretic bacteria, have been shown to cause harm to mammals. Indeed, as mentioned elsewhere, they must be coaxed, cajoled, and pampered to make them work. They comprise, after all, a package engineered by evolution, through millions of years of natural selection, to kill only enough of a narrow range of organisms to survive. Man is able to use them to nullify termite colonies only by repeatedly inoculating specially-designed food sources where the members of a termite colony specifically targeted for elimination have been enticed to feed.

When used in that manner, they are efficient servants, doing man’s bidding without harming non-target organisms. They do their work without exposing the human applicator, or those who live in or at the site where they are used, to collateral risks. This faculty distinguishes these biological agents from practically all of the toxic chemical termiticides employed for termite control.

Unusually Viable EPN Families:

Mermithid EPN parasitize invertebrate hosts in what appears to be an unremarkable fashion while completing certain portions of their life cycle. During the initial parasitic stage the EPN feeds on its host’s fluids. Though this weakens the host, it does not cause death; the insect is subjected — one must suppose — to a degree of discomfort and annoyance by the presence of the nematode, but that is the extent of the parasite’s effect. When conditions are right the nematode transitions from parasite to parasitoid, perforating the host’s body and emerging, producing mortal injuries that kill the host outright. These nematodes were first reported by Aldovandrus in 1623, after they were observed emerging from grasshoppers.

Steinernematids and Heterorhabditids, unlike the Mermithids, kill rather than parasitize insects (though some are capable of parasitizing, or forming symbiotic relationships with, other invertebrates in the absence of suitable insect hosts). Due to their phoretic relationship with a specialized bacterium, their insect hosts are generally killed within hours or days after they invade their host’s bodies. The amount of time between the EPN’s invasion and death of the host varies according to environmental constraints such as, in particular, ambient temperature. At only a few degrees above freezing, host morbidity and death may not occur for weeks, while at room temperature the typical infection is generally lethal in 48 hours or less.

The effect of an attack by Steinernematids and Heterorhabditids on their insect hosts has been likened to that of a “guided missile” (Akhurst, R. J., 1993. “Bacterial symbionts of entomopathogenic nematodes”, CSIRO Publications, East Melbourne). Immediately after entering the insect, the EPN disgorge their phoretic bacteria “warheads”, which multiply and produce (1) a toxin that kills the host, and (2) antibiotics that preserve the host’s cadaver. The EPN feed on the bacteria and use the host’s cadaver as an incubation chamber in which to produce multiple infective juveniles, or IJ’s. Eventually the EPN IJ’s emerge to search for new hosts.

Chance EPN Infections of Termites in Nature:

Termites sometimes suffer chance infections from Steinernematids or Heterorhabditids that are naturally dispersed in ordinary soil. Though such infections often result in the death of the affected termite, the impact on the termite colony itself is generally minor and of limited duration, unless conditions are right for the EPN involved to mount repeated, massive intrusions into the termite colony’s workings. The disabilities that limit the natural virulence of EPN in the wild are well documented. It is usual for those disabilities to be expressed in negative, rather than positive, terms. In general, however, we should be thankful that EPN are so disabled. Nematodes have an indirect influence on most aspects of our existence, yet they have so few direct, observable effects that we are mostly, if not entirely, unaware of their ubiquitous presence. This paradox results from the fact that while these organisms are efficient propagators, they are also rather fragile.

The truths expressed in the following poetic description, though penned by Nathan A. Cobb in 1914, remain undisputed today:

“In short, if all the matter in the universe except the nematodes were swept away, our world would still be dimly recognizable, and if, as disembodied spirits, we could then investigate it, we should find its mountains, hills, vales, rivers, lakes, and oceans represented by a film of nematodes. The location of towns would be decipherable, since for every massing of human beings there would be a corresponding massing of certain nematodes. Trees would still stand in ghostly rows representing our streets and highways. The location of the various plants and animals would still be decipherable, and, had we sufficient knowledge, in many cases even their species could be determined by an examination of their erstwhile nematode parasites.”

Natural Impediments to Virulence in EPN:

Because EPN species are host specific, they can be trusted not to injure non-target organisms. Furthermore, though EPN succeed in infecting many of the insect hosts found in the soil that both organisms inhabit naturally, they are unable, under ordinary conditions, to mount so pernicious an attack that they succeed in eradicating such hosts from that common habitat. As occurs with parasites and parasitoids in general — including those with free-living propagules, i.e., structures like seeds, spores, and juvenile infectives, each capable of giving rise to new, fully functioning organisms in kind — nematode biology developed along lines that militated against untoward virulence:

In nature, EPN-insect interactions involving either the Steinernematidae or Heterorhabditidae, though invariably lethal to the individual insects they manage to infect, typically establish a kind of uneasy equilibrium. Thus the EPN and their insect hosts manage to coexist, though the hosts are somewhat fewer in number than would be the case absent the EPN. While a degree of insect suppression occurs, insect “control”, to the extent that the insects are unable to carry out their characteristic patterns of behavior, does not.

By way of contrast, EPN-insect interactions involving the Steinernematidae or Heterorhabditidae that are bolstered artificially by man, often produce excellent control of many important insect populations. In the past, the subterranean termite has been considered by many authorities to represent a major exception to that rule. Termite and nematode biologists have fretted for years about the difficulties they’ve encountered in efforts to get EPN to work as well in the ground as they do in a Petri dish. Not long ago, one eminent nematologist explained it this way:

“In lab exposures, conducted in Petri plates, host-parasite contact is assured, host escape is impossible, and environmental conditions of temperature, moisture, and light are optimal for infection. In the field, behavioral and environmental barriers come into play and restrict host range. An experimental (i.e. lab-derived) host range should not be confused with field activity. Experimental host ranges can be huge. But in the real world there are barriers that can disrupt the infection process, frustrating control efforts and resulting in a far narrower spectrum of insecticidal activity.”

The above quote is from an Internet article written by Dr. Randy Gaugler, Rutgers University, in an early version of the Internet page now published at Cornell University on biocontrol and nematodes. The language used in later versions of the same web page differs somewhat from the above, perhaps reflecting a positive change in the way academia, or at least Dr. Gaugler, views the use of EPN as agents for termite control.

Many other authorities in the fields of termite and nematode biology have reached similar conclusions. L. H. Ehler, in 1990, pointed out the superior reliability and consistency of chemicals, over nematodes, for insect control, spurring research into ways to improve their performance. Andrzej Bednarek, also in 1990, linked unpredictable nematode efficacy with poor persistence in the soil. J. Curran, in 1993, reported that 90% of the nematodes applied to soil died within a week. Numerous investigations have sought to isolate the various causes of nematode mortality. Some focus on predators and pathogens, others on environmental factors such as temperature, moisture, pH, and the presence or absence of certain chemicals or nutriments.

While reading the material in the remainder of this article, remember the challenges implicit in Dr. Gaugler’s early remarks on this subject. Next, consider carefully the directions taken in the negative reports published elsewhere. To some, including this author, the recognized obstacles listed in these papers suggest, not so much that EPN are unlikely candidates for termite control as that it should be possible to successfully interface EPN with termites in control settings freed of all the cited impediments. That idea set the stage for the investigations discussed in the remainder of this article.

Pest Management with EPN:

Most of the time, pest management objectives require that the targeted insect be (1) exterminated entirely or (2) reduced to population levels that leave them unable to cause damage of economical importance. It is impossible to produce either level of insect control using EPN without first creating unnatural conditions orchestrated and maintained by man. Yet, with an optimum set of artificial conditions, a resourceful user can bolster EPN functionalities to the point that they are able to perform as well as, and in some respects even better than, chemical pesticides. Further, the potential of fine tuning those conditions to the point that they enable EPN to outperform chemical pesticides was significant.

Some user-established sets of artificial conditions work well with EPN against certain target insects. Others fail miserably against one target insect, but work well against others. Certain insects — like subterranean termites — occupy unusual biological niches wherein traditional EPN application techniques are either ineffective or unreliable. For example, in tests using single-locus, inundative applications of EPN to protect homes from large, widespread, vigorous colonies of subterranean termites, the EPN sometimes appeared to fare poorly, though in other cases (using single-locus inundative applications of EPN) they performed to the point of extirpating the subterranean termite colony altogether.

Abortive Experiments with Inundative Termite Treatments:

Under natural conditions, it seems unlikely that repetitious invasions of termite colony workings by large numbers of soil-based EPN occur. However, it has long been theorized that–by using traditional inundative treatment methods–conditions could be artificially produced that protect homes from termite attack. History and custom seem to reinforce this theory, as EPN are successfully applied inundatively as agricultural pesticides, and exterminators and consumers alike have used chemicals and EPN in similar modes against termites since the 1940’s.

Inundative applications of chemical pesticides can be made to cover an entire yard, or less expansively in narrow barrier inundations. Professional termite managers often apply termiticidal chemicals using the latter approach. For years the inundative termiticide of choice was a mixture of the two chlorinated hydrocarbons chlordane and heptachlor.

One inundation, or drench, of a band of soil around a home with a chlordane/heptachlor mixture protects the home from termites for more than fifty years. After all uses for chlordane, heptachlor, and related chemicals were banned by the EPA in 1987, many less-persistent substitutes surfaced. Today, an inundative termiticide is considered acceptable if its lethality, against termites, can be trusted to persist in treated soil for five years or more.

Since certain EPN kill termites, it seems only logical to expect EPN to perform in inundative treatments in more or less the same way as toxic chemical termiticides. Inundating a band of soil with millions of EPN in one or more of the species Steinernema carpocapsae, S. feltiae, or Heterorhabditis bacteriophora, in an unbroken band around a wooden structure, would be expected to produce some degree of control over the termite colonies whose workings traverse the area.

In the 1980’s and 1990’s, Brad Kard, then a termite biologist with the U. S. Department of Agriculture (now professor of urban entomology at Oklahoma State University), tested soil-barriers using chemical termiticides in Mississippi while Roger Gold, professor of urban entomology at Texas A&M University, did the same in Texas. A number of parallel tests were also undertaken using EPN. Dr. Gold supervised many of the latter tests, and all of them failed.

Not one of the documented experiments using EPN in the mode of inundative termiticides passed the crucial test of persistence. A five-year residual — the minimum standard for toxicant-chemical termiticides — was out of the question. Within months after EPN were placed in the soil around wood objects, termites breached the treated band and began feeding on the untreated wood.

On the surface it seems paradoxical that many homeowners claim to succeed in using EPN in inundative applications to protect their homes from termites. Statistics are not available to show how effective these treatments are, but anecdotal reports of successful experiences abound. This contrasts with the poor results of past scientific tests, and the aforementioned list of well-known EPN disabilities that scientists cite as causes behind those dismal results.

All good stewards of the scientific method agree that anecdotal evidence is unreliable. Yet, we must also note that even rigorous scientific tests produce results that require careful interpretation. Furthermore, scientific methods appropriate for one application may be inappropriate and/or inadequate for another. Anecdotal claims should not be considered false merely because they are anecdotal; still, such claims cannot be relied upon until proved in the course of rigorous scientific investigations. The scrupulous scientist constructs experiments, conceived and executed according to accepted procedural and statistical protocols, to remove all possible doubt.

EPN and toxicant-based termiticides work according to divergent, and largely incomparable, modes of action. It follows, therefore, that testing regimens appropriate for one may be unsatisfactory for the other. Scientific tests of toxicant-based termiticide persistence use singular, rather than repetitive, termiticide applications. Toxicant-based termiticides, following the lead of the quintessential historic benchmark, chlordane, are intended to produce effective, persistent barriers against termites with one soil drench event. Under similar conditions, inundative treatments with EPN will almost certainly produce negative results. This begs the question of whether, under other conditions, specifically those better matched to EPN biology, acceptable control can be demonstrated.

Testing EPN on Termites in a Perfect World:

Consider, for a moment, a hypothetical perfect world. We mean by that a world in which every form of bias had been eliminated. Such is the kind of world in which pure science thrives best. The academic setting is naively touted as representing precisely that kind of world, though — admittedly — such worlds rarely exist within the framework of practical reality. This is true even — and some would say, especially — within the most respected academic institutions in the U.S., given the near-ubiquitous dependence of such institutions on funding from industry, and the powerful biases that such funding (and fears attending its loss, should those biases be disrspected) introduces into the functioning of their scientific laboratories.

That caveat aside, one can still be excused for wondering how, within such a hypothetical world, EPN might be properly tested against subterranean termites.

One answer would be that inundative methods would still be investigated, much as history records the abovementioned tests carried out by Brad Kard and Roger Gold. Further, they would likely fail, just as before. But — in a perfect world — such failures would not become EPN’s epitaph. Instead, additional testing, tailored to the unique faculties possessed by nematodes, would ensue. Later tests would take the unusual step of positioning the targeted subterranean termites where they are most vulnerable. Subterranean termites are quite amenable to such positioning, through the use of specialized cellulose-containing canisters placed in the soil where they forage.

Consider once more the list of EPN disabilities that Dr. Gaugler pointed out. Does the existence of such a list, bolstered by the failures observed in inundation experiments, justify striking EPN off the list of potential contenders in the field of termite control? Nothing in Gaugler’s list suggests that nematodes are unsuited for such a role. The first thing he points out, in fact, is how well nematodes do in the petri dish. Why would they not also do well in the field? One wonders why any serious investigator, especially one willing to give EPN a chance, would allow that question to rest unanswered.

The moment inundative methods with EPN produced less than stellar results, alternative methods for applying EPN could and should have been evaluated.  In a perfect world such testing would have been scheduled and executed within a reasonable time period and with the same enthusiasm and coordinated effort as other tests conducted with chemical termiticides.

We don’t live in a perfect world. Scientific research is expensive and, done right, consumes gobs of time. That is one reason it has taken me so long to make significant progress in this field, and I am not even employed in academia, where even non-profit research can use some of the scraps that fall from the for-profit project tables. Question: What would be needed, in today’s economy, to motivate scientists in our universities to pursue further investigations into the potential utility of EPN as biological termiticides?

The fact is, absent obvious tangible benefits, little motivation for such studies exists. Most funding for scientific research on particular kinds of toxicant-based termiticides comes from sources within the industries that produce toxicant-based termiticides. Chemical producers of patented toxicant-based termiticides are blessed with generous margins that can be siphoned to fund extensive, coordinated research that promotes their proprietary molecules. Chemical producers who do not fund such research don’t sell their products, so research scientists in academia quickly learn how to compete for a share of the pie. University labs that do well in this funding arena are able to buy the best equipment, hire the best scientists, and attract the brightest students.

Nematode producers, by comparison, have no special rights over the propagation and marketing of the nematodes they produce, and make do with shoestring budgets that cannot be stretched to cover much, if any, out-of-house scientific research. They cannot offer scientists in academia any of the incentives that the chemical industry can. This leaves a smattering of non-profit research foundations to pick up the slack for them. Non-profit research, though, is less coordinated, less intensive, and rarely focuses on promoting a particular method or agent.

Such, as they say, is life. Now, at this point I need to explain why we are even discussing this. It isn’t because I like to gripe. The reason lies in quite another direction. It happens that, once university professors and their research staffs decide to dismiss a product or method because they say “it doesn’t work”, their published opinions carry a lot of weight in the real world, even if the basis for such remarks has little or nothing to do with true, objective, evaluation.

Today it is possible to find, on the Internet, a multitude of dismissive statements from academia and industry that purport to establish that EPN are incapable of performing in the field of termite control. The material presented here effectively counters those statements and shows that they are incorrect. In fact, in the following it will be shown that EPN are capable of performing in a stellar manner at termite control. In particular, monitored devices that intercept and concentrate a significant fraction of the subterranean termites associated with a given subterranean termite colony, set the stage for inoculating those termites with EPN. As early as the mid-1990’s, I suspected that if the devices involved could be optimized for EPN inoculation purposes, they would potentiate a residual agency of termite control comparable or superior to that of every chemical termiticide presently on the market. All my field studies since that time, conducted in five of the six major biotic provinces of Texas (excluding only the Chihuahuan biotic province, in the far southwestern corner of the state), have since shown that suspicion to be correct.

An Unwarranted Dismissal:

Based on results derived from single-application inundative testing, most researchers in nematology, entomology, and the pest management industry concluded that EPN could not be relied upon as biological termiticides. That conclusion was correct with respect to the limited mode of single-application inundative treatments. However, the suitability of other modes of treatment remained to be explored, but few have been willing to take up that torch.

It will be shown, in the material that follows, that blanket dismissals of EPN for termite control are unsupportable. Yet, and this is not a surprise, few scientists have advanced serious challenges to them. Now we must ask a crucial question: does it even matter? If EPN really don’t offer anything special to the field of termite control beyond merely being efficacious, in some ho-hum sort of way, the answer might be a resounding “No.” Are EPN special? Should any of us care if they are used, or made available for us to use for termite control in place of other candidates? It seems clear that is the next question we need to address and answer.

Why Use EPN for Termite Control?

Below are some of the reasons why I believe entomopathogenic nematodes (EPN) should be used to control subterranean termites:

1. Because they can: EPN were made to kill termites.

2. EPN are safer than toxicant-based termiticides.

3. Used intelligently, EPN are inexpensive termiticides.

4. EPN are effective. They do what they need to do, nothing more, nothing less.

Natural, Minimum Risk Pesticides are “Known Quantities”…

Some will argue that EPN were not made to kill termites, but in these pages I prove they are not only capable, but are well qualified for that task. Beyond that, most agree that EPN are safer, for applicators and consumers alike, than any of the exotic termiticides now on the market. All who have studied the question in depth know, often from personal experience, how safe natural, minimum risk pesticides are when contrasted with exotic toxicants, In particular, toxicants belonging to new classes of chemistry carry risks to mammalian biology that may not be understood for decades or longer. With the exotics, it is most often what we don’t know that is most troublesome. Natural, minimum risk pesticides have been around for so long, and have been used as pesticides in so many ways, that their effects on humans and other mammals are well-known and documented.

As pointed out elsewhere in these pages, it is logical that EPN be used in close association with other natural, minimum risk pesticides. Such associations tend to achieve the best results against termiticidal agents. It makes sense, therefore, to discuss the natural, minimum risk pesticides that are available to work alongside EPN.

The U.S. Environmental Protection Agency (EPA) exempts entomopathogenic nematodes, and certain other natural, minimum risk pesticides from regulation. By itself, that does not make such materials absolutely safe. Still, a user of such products can be certain that their toxicology profiles contain no red flags. Few materials are so honored. Besides EPN, the list of natural, minimum risk insecticidal and termiticidal pesticides that share this distinction contains, at present, only 31 products.

40 CFR ‘ 152.25(f), notice 2000-6, lists the 31 natural, minimum risk pesticides, many with known termiticidal properties, that are exempt from the requirements of the Federal Insecticide, Fungicide, and Rodenticide Act (FIFRA). The list includes a mineral, sodium chloride–or common table salt, whose mineral designation is halite; it happens that sodium chloride is used quite successfully as a termiticide in some locales today. This list also contains a metal, zinc, when made into strips that are used as a fungistat to prevent and control algae and fungi on roofs and buildings.

Also on the list are botanical extracts and by-products, including malic acid (the source of sourness in green apples); cedar oil (which has exceptional termiticidal qualities); citric acid; citronella and citronella oil; cloves, clove oil, and eugenol (an extract of clove oil and a number of other botanicals, also with exceptional termiticidal qualities); corn gluten meal; garlic and garlic oil; geraniol; geranium oil; mint and mint oil, & peppermint and peppermint oil (the mint oils are also potent termiticides); 2-Phenethyl propionate (a natural fungicide and pesticide derived from peanut oil); potassium sorbate (a potassium salt of sorbic acid, which is naturally found in the unripe berries of plants in the genus Sorbus); rosemary and rosemary oil (also potent termiticides); sodium lauryl sulfate (a foaming agent and surfactant naturally derived from coconut or palm kernel oil); and thyme and thyme oil (again, with good termiticidal qualities).

These natural minerals, botanical byproducts, and plant extracts have more or less existed with and evolved alongside mammals throughout history and beyond. Their effects on mammalian biology are both limited and well-known. Some, including common table salt, cannot be consumed by mammals in excess without causing easily recognizable, transitory health problems. For others, ingestion of small amounts of, or direct dermal contact with them produces either a beneficial effect on mammals (eugenol is used as a local anesthetic in dentistry) or no noticeable effect at all.

Some important, safe and natural pesticides that are not on the 40 CFR ‘ 152.25(f), notice 2000-6 list–and that are not, therefore, exempt from EPA registration–include boric acid, borax (sodium tetraborate, with four boron atoms/molecule), and the enriched borate salt disodium octaborate tetrahydrate, which has eight boron atoms/molecule). These natural pesticides have safety profiles comparable to those of certain exempt minimum risk pesticides (e.g., sodium chloride). To many, myself included, It seems appropriate to rank the borates alongside the exempt minimum risk pesticides, even though their uses are strictly regulated by the EPA, and the instructions on their EPA-mandated labels must be followed to the letter.

Practically all of the natural materials described above are capable of serving as proximal, temporal, and spatial synergists and adjuvants with one another and with EPN. Lab and field experiments show that treatment programs that use EPN in combination with certain natural mineral and plant-oil termiticides achieve superior results to regimes that use only one of these materials in isolation.

Exotic Pesticides Raise Questions We cannot Answer…

Man-made termiticidal agents are, in general, rightly classed as exotics; this word denotes something unnatural, i.e., a material whose essential molecular structure is not now, and was not in the past, commonly found in the environment. Long-term risks posed by exotic termiticides cannot be ruled out, even if rigorous testing suggests the absence or paucity of short-term risks. That is why it is necessary to place baits containing exotic toxicants in child-and-pet-resistant containers. Yet, no matter how hard man tries to do so, the leap from “child-resistant” to “child-proof” remains a dream. Never mind, of course, the leap from “pet-resistant” to “pet-proof”.

EPN used for bio-control purposes, on the other hand, pose no danger to mammals:

“Since the first use of the entomopathogenic nematode Steinernema glaseri against the white grub Popillia japonica in New Jersey (USA) (Glaser and Farrell, 1935), not even inferior damages or hazards caused by the use of EPN to the environment have been recorded*. The use of EPN is safe for the user. EPN and their associated bacteria cause no detrimental effect to mammals or plants (Poinar et al., 1982; Boemare et al., 1996; Bathon, 1996; Akhurst and Smith, 2002).” Dr. Ralf-Udo Ehlers Christian-Albrechts-Universität, Kiel, Germany, Safety and regulation of entomopathogenic nematodes. [*Emphasis added].

EPN deserve serious attention, therefore, on the basis of intrinsic safety alone. Today, our knowledge of EPN biology contains much that is positive with respect to their suitability as bio-control agents. In fact, every one of their well-known natural disabilities is, in reality, a positive attribute when viewed vis-à-vis the greater issue of mammalian safety. That is why I believe EPN should be given every chance to serve mankind in the field of termite control.

Giving EPN a Chance:

EPN, when given a chance, are hardy organisms with amazing talents. Many species appear well-adapted for controlling insects like termites. Furthermore, the more they are evaluated in the field, the more capable they appear. One by one, their much-touted disabilities cease to be seen as problems and are viewed instead as beneficial qualities that make them safer around mammals. Some of their perceived weaknesses don’t even exist, when examined with care.

For example, EPN are able to persist in the soil for decades if conditions are right. When used in EPN-optimized termite interceptors they can be made to persist (through repetitive inoculations), in the interceptors and wherever they are transported by infected termites into the workings of the termite colony, at least as long as the termites they are intended to exterminate, and that is long enough. Still, the concept that EPN are weaklings, susceptible to every environmental condition imaginable, is legendary. One popular website confidently, but incorrectly, asserts that

“(EPN) move no more than 18 inches in their lifetime on their own and will die in temperatures of over 70 degrees…”

It may be possible to find EPN that can be described that way, but none of the EPN we have worked with are so handicapped. It is true that some EPN, known as ambushers, do not move far once they are inoculated, but that is merely a result of the way they behave in nature (ambushers wait for prey to come to them, while cruisers actively seek prey out). A user that wants EPN that tend to move around will use a cruiser species, not one that prefers to ambush its prey. Many EPN, whether ambushers or cruisers by nature, are adept at hitching rides on or inside a variety of commensal or symbiotic hosts such as earthworms and other soil-based organisms, and thereby travel long distances. Most survive when subjected to temperatures in the high 80’s and low 90’s.

The range of environmental conditions suitable for EPN survival expands with each new spate of pure research, and with discoveries of new EPN species. That range grows even wider as investigators document the behavior of EPN as temporary parasites of earthworms and other, non-insect invertebrates.

Effective Soil Inundation Treatments with EPN

Soil inundation with entomopathogenic nematodes (EPN) to control fleas and/or grubs is an effective, safe means of insect control. If you remember the material on the preceding pages, you may now be scratching your head, saying “I thought Jerry said inundative treatments using EPN are not a good idea.” Well, I did say that, but with a caveat. Stick with me, now. I’ll try to make sense out of this as we go along.

Inundative treatments with EPN for termite control are, in my judgment, practically worthless. But the same is not true of EPN inundations for certain other pests, like fleas. I often treat yards for fleas in this manner, always with excellent results. One of the most satisfying aspects of these kinds of inundative EPN treatments is knowing that the EPN I inundate the soil with will attack only the larval pests in that soil, but will leave most, if not all, of the beneficial organisms alone.

But I never use EPN in inundative treatments to control subterranean termites. My reasons for this distinction should become clear in the material that follows. In summary, though, inundation for termite control is wasteful; it is not an intelligent way to use these incredible, capable organisms.

Using Toxicant Pesticides for Broad-Spectrum Soil Inundation Treatments:

Let’s put EPN aside for a moment and take a look at the typical way most pest control companies treat pests. They use broad-spectrum toxicant pesticides, as a general rule, despite the fact that using such products to inundate the soil, regardless of the target insect involved, is both the least precise and the least economical method imaginable. See? It is probably never a good idea to inundate soil with broad spectrum toxicants, The technique violates a fundamental rule of logic, because it causes more pest problems than it fixes. Why? Because broad spectrum pesticides have significant collateral effects that extend far beyond the original treatment objectives.

Inundating whole yards, or even narrow strips within them, with residual, broad-spectrum, toxicant pesticides is akin to the practice–in military warfare–of laying a huge number of landmines around a fortress or a weak place in a border area to defend against a few small and scattered interlopers. Yes, I know such treatments are carried out by pest control companies every day in almost every community across America, usually at the express insistence of their customers. And yes, for a short time at least, those treatments successfully wipe out important target pests like fleas, ticks, grubs, etc., and make pest control customers happy. But, to be honest, the only reason these customers are happy is that they don’t realize the total cost they are paying for those treatments.

Whole-yard inundations with broad-spectrum toxicant pesticides are not only inefficient, but counterproductive. As with the landmines used in warfare, only a few intended targets are destroyed by the killing agent, while causing collateral (in other words, secondary, entirely unintentional) destruction of multitudes of other organisms–many of which have a beneficial role in the environment. I will have more to say about these beneficial organisms in a moment.

Applying Toxicants in Soil Inundation Treatments for Termite Control

What about toxicant termiticides for inundation treatments? A number of toxicant termiticides are now labeled to be used for soil inundation, and a few have broad labels that include practically every other insect pest known to annoy or afflict man as well. Some show high persistence in the soil, lasting for months or years following the application.

Often, when the target pest is the subterranean termite, the applicator will dig a trench around the perimeter of the structure to be treated and will then inundate the trench and the back-fill used to fill it back up. This produces a barrier to termites that attempt to tunnel through the toxicant-laced soil. Some are repellent and turn termites away, while others are not repellent and the termites, unaware they are being intoxicated, keep on digging until they die. Toxicant termiticides applied in this manner can persist for many years. Most remain toxic to termites–and all the other insects inhabiting the same soil–for at least five years, and some as much as 15-25 years or longer.

Though some termites are killed, as they tunnel into these toxicant-laced barrier strips, most members of the termite colony will continue to thrive without injury, regardless of the kind of termiticide used. This is because they don’t happen to travel in the workings that traverse the inundated portions of the colony’s extent. Of course, it is possible to eliminate termite colonies in whole yards by inundating the entire yard with these broad-band termiticides, but, fortunately, that is too costly a practice for most consumers’ pocketbooks. As we shall see, it is also too costly from another angle–the slaughter of all the beneficial insects in the yard–as well.

The Importance of Beneficial Insects in the Environment

I realize the value of beneficial insects will not seem important to those of you who do not understand how they work. But once the beneficial insects are destroyed by broad-spectrum toxicant treatments, a whole list of other pest insects–such as acrobat ants, carpenter ants, puss caterpillars and so on–that are usually kept under control by a host of beneficial insects that prey on them, quickly grow more numerous.

To many people, that merely signals the need for another application of broad-band toxicant insecticides, i.e., more of a bad thing, in what soon becomes a never-ending cycle. Fortunately, it isn’t hard to get a grip on why beneficial organisms really matter. Once your understanding on this subject improves, it will be hard for you to justify using broad-band toxicants in the future. And once you stop that wasteful practice and begin using rational, minimum risk pesticides that target specific organisms, your life will improve in ways you cannot imagine.

The Fallacy of Broad-Spectrum Toxicant Pesticide Treatments:

But First: A Lesson Taught by Fire Ants and Food Competition Beneficials

Over the past thirty years, I’ve been taught many good lesson in the importance of beneficial organisms. Sometimes those “beneficials” attain that status because they prey on important pests, but at other times they are important merely because they compete with such pests for food. At other times, they become valuable because they prey on symbiotic organisms that the pest insects need in order to thrive.

Fifteen years ago, I was called to a nursing home in north-central Texas where fire ants had taken over the landscaping. The pest management company responsible for fire ant control at that site had been spraying broad-spectrum pesticides throughout the facility’s landscaping for the previous two years. Now, suddenly, fire ants were out of control and the nursing home was in a lot of trouble. The Texas Department of Health, after being called out by the family of one of the nursing home residents, had cited the nursing home with an “immediate jeopardy” violation because fire ants were stinging residents in the yard and getting into their beds. Fines levied by that agency, for this single violation exceeded $50,000 before the charge of “immediate jeopardy” was finally lifted (the fine was levied on a daily basis, even though the agency would not revisit the facility for at least 30 days to conduct an inspection that, when passed, allowed the violation to be cleared).

Let’s examine why this fire-ants-from-hades incident occurred, because it doesn’t follow the usual path you might expect to see with the destruction of ordinary beneficial insects. Here, rather than beneficial predators, it was the beneficial food competitors that were being killed. You see, until beneficial fire ant predators were recently introduced, imported fire ants had no serious natural predators to worry about in the United States. But they still had to compete with other insects for the available food supply. In effect, that competition kept their numbers in check. In an environment that does not have any food competitors, but that contains plenty of food, the fire ant population can explode from a handful to literally tens of thousands overnight.

At this nursing home, the pest control company had destroyed all the fire ant’s food competitors, so whenever a new fire ant colony dropped in, it quickly grew huge and strong on the bounty of food it found there. The other pest control company had given up on trying to control fire ants on the property because no matter how much they sprayed, the fire ants managed to thrive. Evidently they didn’t believe anything but spraying was worth doing.

I was able to bring the fire ants under control, within days, using a granular bait that targeted fire ants specifically and left all the other insects alone. Over the next few months, as the effects of the other company’s past spraying wore off, the usual food competitor insects slowly returned. Before long, I was able to significantly reduce the amount of granular bait necessary to keep the fire ants in check because the other insects were doing their part in the process as well.

Next: A Lesson Taught by the Puss Caterpillar’s Natural Predators:

Another lesson was taught to me more recently by the puss caterpillar (Megalopyge opercularis). Its common name derives from its similarity, in appearance, to a tiny, furry kitten. Though this is one nasty bug, it looks so soft and cuddly that kids and adults alike just want to pick it up and play with it. That’s not a good idea, because one contact with this stinging caterpillar produces what many describe as the most painful sting ever. The pain is so bad, and so pervasive–it quickly migrates to the lymph glands in the armpit or groin, then to the chest–that most who get stung end up in a hospital Emergency Room convinced they are about to die. One puss caterpillar sting can easily lead to many hours of lost productivity, and medical bills that can run upwards of $1,000 or more.

I began keeping track of outbreaks of puss caterpillars (CLICK HERE to see my articles on this serious insect pest’s natural predators) throughout the United states in 2002, after encountering some puss caterpillars infesting yaupon holly–a favorite food of theirs–in the landscaping of two Texas nursing facilities, one in Georgetown, the other in Temple. I discovered, first hand, that aggressive treatments using broad-band toxicant pesticides only led to fresh outbreaks, in the same place, later on. Further, in every case reported to me from all over the United States by others, where aggressive, broad-band pesticidal treatments were used, the caterpillars returned, usually in larger numbers than before.

Curiously, though, when the caterpillars were removed by hand (using heavy rubber gloves), or flushed out of bushes with streams of water, their infestations lasted only one season and never returned.

It was obvious to me that, in most cases, it is impossible to collect all of the caterpillars using manual pickup methods, and that many won’t be flushed out of the foliage where they are feeding using a simple steam of water. In fact, these natural and manual processes allow significant numbers of puss caterpillars to survive and pupate. In the normal process, the pupa emerges later as a moth, to resume the life cycle of the species. Yet both of these methods, when not supplemented with pesticides of any kind, work extremely well: the puss caterpillar infestation is shortened, and successive infestations are not observed in the next season.

When aggressive spraying with toxicant pesticides is done, a few caterpillars won’t be killed, just like when the other methods listed above are used. But there is one essential difference. When using manual and/or hydraulic removal, none of the beneficial insects that prey on Megalopyge opercularis are harmed, so the beneficial predators of the puss caterpillar remain ready and able to take up the slack. These beneficial insects work so efficiently, in fact, that they prevent another outbreak from taking place even when large numbers of caterpillars have to be killed.

Not so when pesticides are used. Most or all the beneficials are killed by the pesticidal chemicals, along with some, but not all, of the caterpillars. Then, because the small number of caterpillars that survive are not bothered by their normal insect predators, they often return the next season in even larger numbers than previously.

We’ve gotten slightly off-track from the main topic, but I felt it important to at least address the problems that come from using pesticides in any treatment program. Now, though, we should wander back to the question of using soil inundation for termite control.

Chronicity vs. Acuteness:

The effects of traditional, inundative treatments with toxicant pesticides for termite control are intended, by the very nature of the treatment regimen, to be long-lasting. The toxicants involved are supposed to render the soil they treat chronically toxic. The best agents to use with such treatments, therefore, emphasize chronicity over acuteness.

EPN used for termite control do not produce a chronic toxicity of the soil in which they are placed (a definite positive for EPN). Instead, they possess a potentially lethal symbiotic bacterium that is only released once the EPN enters the body of a target organism. Each EPN is very organism-specific, not only because of the habits and preferences of the EPN itself, but also because its symbiotic bacteria won’t dislodge itself and multiply inside just any old organism. The pH, for example, has to be within a very narrow range, and other chemical factors have to cooperate as well.

This symbiotic bacterium, though, once it is released into the blood supply of a suitable organism, produces an acute infection that kills the organism within hours. Once death occurs, the dead insect’s cadaver becomes a perfect incubator for new EPN, because the symbiotic bacteria also produce antibiotics that keep the cadaver fresh by killing microbes that cause normal putrefaction.

To the termites who pass by, one of their workers has died, but because the worker’s body doesn’t decay they treat it like a live one. Days go by, and still nothing happens, but finally, sometime later. the cadaver ruptures and a fresh batch of EPN emerges. Again, the termites passing by do not notice, but if they get too close they might feel an EPN crawl into their mouth or go up the orifice at the other end of their body. When that happens, the termite will soon get sick and die, as the infection cycle repeats itself.

The mechanism of control with EPN involves a subset of the workers in the termite colony. That subset becomes infected by the EPN and quickly succumbs from an acute infection. To the uninfected members of the colony things continue as before, but with fewer of their workers doing the work, but the reduction in workers isn’t alarming. Until the next infection cycle starts, nothing else happens. In fact, the onset of a new infection cycle isn’t of any real concern because the process is so low-key. Periodically, dead termites crack open and fresh EPN do what they do to begin the process of removing more workers from the colony’s roster, but that’s all they do. Over time, with repeated inoculations of EPN, the termite colony is reduced to a fraction of its former numbers. Eventually, the colony ceases to exist.

None of this involves chronic poisoning. The EPN do have to find a way into the termite colony, however, so they can come into intimate contact with the termite workers, or none of their useful termite-destroying capabilities will amount to a hill of beans. And because inundation techniques don’t do anything to get the EPN in the door, guess what? They don’t do well when used that way.

Inundation methods have no utility with EPN, when used for termite control, because inundation emphasizes acuteness over chronicity. As a result, little has been gained by testing EPN against termites using inundation techniques, even though that is just about the only way such testing has been done in the field. That such tests even took place illustrates a widespread lack of knowledge about EPN that existed then and continues to exist today within the field of termite biology.

Historic Trends in Academic Research:

With history as our guide, such a lack of knowledge is understandable. The state of termite biology before 1987 was abysmal because, as long as chlordane reigned supreme, industrial sources of funding for research into alternative termiticides was scarce. Since most academic research in termite control was (and is) funded by the chemical pesticide industry, the direction that industry takes tends to define the direction of university research.

By 1987, when chlordane and its related chemistries were banned for termite use, academic knowledge was accelerating at an astounding pace. Huge outlays of new funding poured into termite biology projects. A similar explosion in our knowledge about EPN-termite interactions is needed, yet almost no sources of funding are available.

It is easy to see why. You may be able to patent a novel species of EPN, but you will likely fail to prevent others from culturing your patented species in their own labs, after they buy a small seed quantity from you. They will then be able to cut you out of any future profits you might otherwise reap. Here is what that means: since future suppliers of even novel species of EPN cannot expect to earn profits comparable to those enjoyed by manufacturers of toxicant chemical pesticides, they will have to settle for less, just as present EPN suppliers have to do. The question is, will such profit margins be enough to sustain a viable EPN market and fund the research needed to take that market forward?

That is a two part question, and the answer to the first part is yes, but the answer to the second part is probably no. Sustainable markets for EPN already exist, and there is good cause to believe they will remain that way. But it is a fact that the number of studies carried out since 1987 on EPN-termite associations pales in comparison with the mountain of studies conducted since 1993 on the effects of a single termiticide molecule, namely hexaflumuron; is seems likely that the state of EPN research will remain at that dismal level into the future, as well.

Regardless of the limited funding for academic research into EPN suitability for termite control, a smattering of studies continues to be done, and a number of non-profit and for-profit research projects are underway at companies like mine whose focus is on biological control methods. And, despite the fact that we are forced to do our own research, using our own in-house funds, we still manage to achieve results.

At EntomoBiotics Inc. our lab and field studies have shown that even though inundation is not a viable way to apply EPN for termite control, other methods hold great promise. Yes, EPN are made to kill termites, they just weren’t made to kill them using soil inundation methods. There are other ways that work. One of these in particular — inoculation of EPN into special termite interceptors — is poised to make EPN fully capable of eliminating termite colonies using natural methods without the use of exotic toxicants.

Where Inundation Failed, Inoculation Succeeds:

Before going into detail on how EPN work as inoculants in specialized termite interceptors, we need to detour again. This time, we will look into how the toxicant termiticide hexaflumuron is used to inoculate termite colonies and eliminate them.

No inundative termite treatment method — whether chemical or EPN — reliably eliminates termite colonies. However, placements of minute quantities of termite control agents, within the confines of a termite colony’s workings, are now known to be capable of utterly destroying the affected termite colony.

The exotic toxicant hexaflumuron, a chitin synthesis inhibitor, was the first termiticidal bait ingredient to demonstrate termite colony elimination. Similar molecules, including the chitin synthesis inhibitors diflubenzuron, lufenuron, et al., have since been proven to succeed in that role as well. These chemicals, when placed inside bait stations inserted in the soil actively foraged by a termite colony, intercept the colony’s foraging workers by enticing them to consume minute amounts of the interceptor’s bait before reentering the colony’s workings.

The termite workers mark the pathways to the bait station with a pheromone to show that it contains a suitable food source, and this serves to attract other workers to the station as well. The workers that feed in the bait stations later pass fecal matter contaminated with the active ingredient in the bait. Subterranean termites practice proctodeal feeding, or recycling each other’s feces in what amounts to an efficient method of continuously replenishing the gut fauna of all the termites in the colony. We may find the mechanics of that method disgusting, but termites do it as a matter of absolute necessity: proctodeal feeding, also termed proctodeal trophyllaxis, serves to insure the maximum utility of nutritional matter by maintaining the efficient distribution of gut fauna throughout the termite colony.

The only downside of the termite’s practice of proctorial feeding is that if one or more termites in the colony consume(s) a lethal chemical that is later passed in its anal secretions, that lethal chemical has the potential to also become well distributed within the colony. Hexaflumuron meets those criteria, and termites that feed in bait stations containing hexaflumuron soon begin to leave a trail of hexaflumuron-laced termite feces that effectively pass the hexaflumuron molecule throughout the colony’s workings, so that even the termites that have not fed in the bait stations containing hexaflumuron bait are exposed to it secondarily. Sometime later, as each of the termites contaminated with the hexaflumuron molecule attempts to molt, their new skins are prevented from synthesizing while, coincidentally, their old skins remain unnaturally rigid. The unfortunately immobile termites soon die. Over time, enough members of the termite colony become dosed with the toxicant to destroy the colony completely.

Inoculation in Action:

Placing something where it can grow or reproduce is referred to as an inoculation. The placement of an exotic toxicant in a bait station, so that it will be spread to other parts of a termite colony by the termites that feed there, is one kind of inoculation. Even though hexaflumuron does not grow or reproduce in the termite colony, its effects are transmitted far from the bait station where it is first ingested. The net effect, therefore, is the same as if it did grow or reproduce. Its range of effect is extended beyond the small placement of the bait station in the soil. The bait station itself is a kind of termite interceptor, in that it serves as a feeding node for the termite colony. The feeding termites do not remain in the termite interceptor any longer than necessary to acquire a meal, and when that interval has passed, the termites exit the interceptor and begin the trek to the next food source in their “list” of places to eat.

The concept of using termite interceptors to inoculate termite colonies with toxicants is powerful. It was first applied by Glen Esenther and Ray Beal in the 1960’s, and is discussed more thoroughly in my article on termite interception. During the 1990’s a variety of devices, most designated as bait stations but all serving as termite interceptors, were designed for use with various toxicant inoculums. As my work in EPN/Termite research developed, it became clear that the same basic concept should work for EPN, too.

Inoculations with EPN:

Ordinary termite interceptors designed for chemical toxicants are unsuitable for EPN inoculations. To give EPN a chance, conditions in the interceptor must be optimized, as much for EPN propagation as for termite attraction. With an EPN-optimized interceptor, the EPN inoculum and their phoretic bacteria, on infecting the termites within a termite colony, should become what Dr. J. Kenneth Grace envisioned as the ideal microbial termite control. As noted elsewhere, he wrote that such an ideal microbial agent would work as “a self-replicating time-bomb, akin to a computer virus” (J. Kenneth Grace, Sociobiology Vol. 41A, 2003, in an article entitled Approaches to Biological Control of Termites.)

Thus, instead of randomly distributed landmines with delayed fuses (e.g., using inundative techniques), the “time-bombs” Dr. Grace mentioned are planted in specific locations, in the precise number needed, and only inside specific kinds of desirable target organisms. When the “time-bombs” go off, however, the result is not an explosion that damages unintended targets nearby, but is more akin to the opening of a gate, which frees the agents — from the case in which their clandestine development took place — so they can disperse and attack new target organisms with the same precision as their progenitors.

In actuality, EPN-infected termites behave like mobile, time-regulated misting devices. Outwardly quiescent, they travel far from the interceptor, into the termite colony’s workings where they die from the EPN infection. Because the phoretic bacteria that kill them also produce antibiotics that prevent the termite cadavers from putrefying, uninfected termites do not avoid the cadavers, or wall them off from the rest of the colony.

The dead termites lay dormant and innocuous until, without warning, they fracture and new EPN IJ’s emerge. And because they are restricted to the workings of the termite colony, the IJ’s have almost no opportunity to attack non-target organisms. Confined within the termite colony superorganism, the EPN easily find new termites to infect, because they emerge within active tunnels that are teeming with termites that are traveling through to other locations in the termite colony.

Termites that become infected with EPN inside an EPN-optimized termite interceptor perform much like those coated with groomable toxicants. They soon leave the interceptor and carry their potent termiticidal baggage deep into the termite colony’s workings. The process is similar to the program developed by Dr. Timothy Myles, which he called Trap-Treat-Release (TTR), but without the laborious manual operations that accompany that program.

Subterranean termites use ingenious methods to compensate for the often less-than-ideal environments where their cellulose-based food sources are found. If temperature or humidity is too high or too low, they tunnel to more congenial places nearby and pipe that air and moisture over so they can keep on feeding. Wood that is too dry gets a coat of mud, a splash of the termite worker’s secretions, and as many drops of water — carried in from the nearest water source — as needed to raise humidity to an acceptable level.

Wherever they go, termite workers build and maintain a mud wall that encapsulates the colony on all sides, regulating its environment and eliminating undesirable fungi, bacteria, predators, and parasites. That’s one reason why the subterranean termite colony, in combination with its system of workings, is often referred to as a superorganism. The mud walls of the tubes and enclosures that encapsulate the colony are like an animal’s skin. They form a wrapping that, if visible to the naked eye, would perfectly define the extent of the colony’s meanderings.

Like skin, the workings of the termite colony protect vital interior constituents, by regulating humidity and temperature and excluding exterior contaminants. Subterranean termite workers, if forced to venture outside this skin, soon die from exposure. Inside it, they live relatively long and healthy lives. Some of the live termite specimens in my laboratory collections are over nine years old, and it is likely that a few of the termites that were alive when I first collected those specimens are still alive today. Termite queens are capable of living twenty years and more.

Termites Actually Help EPN Do Their Job…

The meticulous way termites regulate environmental conditions within their workings is similar to the way we regulate the interiors of our homes and laboratories. Ironically, this hard work, which is necessary for their survival, would also help entomopathogenic nematodes (EPN) infect and kill them, if they could only get inside the colony’s confines. That is one of the reasons why, in lab experiments with EPN, termites fare poorly (none survive, in fact, when under attack by EPN). We easily see why: in the lab, where temperature and humidity are kept at levels optimum for termites, they are also optimum for EPN. Termites and EPN share a need to live within a narrow range of temperature and humidity. Fortuitously for those of us who seek to use EPN for termite control, the range that works for one also works for the other.

But Only Inside Their Workings:

Field experiments conducted by major universities, however, failed to demonstrate that EPN could control subterranean termites in the soil. In each of those experiments, termites did well while the EPN fared poorly. Why? The field tests used soil inundation techniques: EPN were poured out, over the soil, outside the workings of the termite colony, and had to find a way inside quickly, to take advantage of the air-conditioned environment. Termites build their colony’s skin as a well-constructed fortress, and getting inside is so difficult that few EPN — if any — succeed.

Once inside, of course, an EPN should manage to perform as efficiently at killing termites as in the typical laboratory experiment that used petri dishes in controlled environments. Better, even, because the close confines found in actual termite workings let EPN gain intimate contact with termite workers right away, while petri dishes give the workers lots of wiggle room.

The Challenge: Get The EPN Inside

This shows how imperative it is for EPN destined for termite control to break through the skin of the termite colony as soon as they are applied. Obviously, inundating the soil with EPN will not do that. I also discovered that plowing up the soil, to assist EPN that have been applied inundatively, fails as well; individual termites immersed in an amorphous soil medium instinctively construct tunnels with hardened walls that they can defend, and they do it quickly enough to stymie their natural enemies, including EPN.

What is needed is (1) a well-defined weak spot in the colony’s skin where, coincidentally, (2) termite workers are known to mass within (3) a confined space, whose constituent parts (4) are arranged to facilitate EPN infections of termites, and that (5) enable continual infections of newcomer termites to take place as well.

These five basic criteria describe, in a nutshell, the design features contained in the biologically-optimized termite interceptor developed at EntomoBiotics Inc.

An EPN-Optimized Termite Interceptor

The termite interceptor we developed for EPN inoculations becomes an integral part of the termite colony’s superorganism as soon as termites begin feeding inside it. Termites incorporate all such interceptors that they find into their cellulose feeding circuit. This circuit consists of food sources and pathways, or transit tunnels, over which they travel in a closed loop. Individual termites take a meal in each food source they come to, then trek to the next one while their gut fauna digests the food from the last meal.

Each food source serves as a feed station where hungry termites congregate in relatively large numbers while ingesting a fresh cellulose load. In any active feeding station thousands of worker termites will be found at any single moment in time, all packed into a relatively small, three dimensional feeding area. Inoculating such a feeding station with EPN could result in massive, simultaneous EPN infections, with consequences that would soon reverberate throughout the termite colony.

The termite interceptor developed by EntomoBiotics Inc. allows EPN to infect the termites inside it soon after inoculation takes place. It does this by incorporating design features that defeat the ordinarily robust integrity of the termite colony’s skin, so that each inoculation of EPN is able to immediately break into the termite colony’s workings en masse. Few or none of the termite workers feeding inside this interceptor at time of inoculation will be able to escape infection, and all will shortly depart, en route to their next food source. This interceptor then continues to harbor residual EPN, which proceed to infect newly arriving termites soon after they arrive.

This process, fortified with periodic inoculations of fresh EPN into the interceptors by the user, maintains a continuous and cyclic infection process within the termite colony.

Each infected termite becomes a ticking, time-actuated release valve. Generally, within one or two days or longer–depending on temperature and other conditions, the interval can be much longer– the termite dies. Because the cadaver doesn’t putrefy, uninfected termites ignore it. Evidence of complex nematode avoidance responses by subterranean termites is sparse to nonexistent, especially with respect to the most common termite species, while evidence that termites ignore, or are attracted to and even cannibalize cadavers infected with EPN, is abundant.

EPN/Termite Interactions

Contrary to certain vague references found in the literature, I have observed no evidence that uninfected termites take steps to wall off EPN-infected members, alive or dead, from the rest of the superorganism. Termites often wall off cadavers of nest mates infected with white or green muscardine disease (a fungal infection), but the practice seems not to extend to cadavers infected with EPN. Personal observations of termite workers, soldiers, and various kinds of reproductives, all in the genus Reticulitermes (whose species are the most destructive subterranean termites in North American), have failed to demonstrate any complex avoidance responses in the presence of nematodes. Researchers working with Formosan termites have noticed the same lack of complex avoidance responses by members of that species to the presence of EPN, which suggests that EPN wil do well against these termites as well.

Simple avoidance responses, such as acting annoyed and moving a short distance away when a nematode touches the termite’s cuticle with its anterior or posterior extremity, are usually observed when termites find EPN nearby. These kinds of responses, however, are not effective at preventing EPN infections. We see this in petri dish experiments, where the termites have lots of room to move about while placing some distance between themselves and the annoying probing of the EPN. Inside the workings of a termite colony, where the narrow confines of each subterranean tunnel work to bring termites and EPN even more intimately together, one must expect the opportunities for the EPN to gain entry to a given termite’s body to increase.

Generally, three to five days after the infected termite dies–temperature and other factors vary this interval considerably, up to and beyond twenty days–the termite cadaver bursts open, and tens or hundreds of EPN IJ’s, along with larger adult male and female EPN, begin to emerge. Afterward, additional IJ’s continue to be produced within the termite cadaver, emerging in a slow procession for 20 to 40 more days. As the IJ’s that emerge from the termite cadaver invade the bodies of, termites that pass by, a new infection cycle begins, in a self-replicating cascade of re-infections within the superorganism.

As figure 1, at the head of this article, shows, the size of the EPN IJ is small, compared to the size of the average subterranean termite worker. The danger of immediate mechanical injury to the interior anatomy of the termite worker, by even multiple invasions of EPN, is low. Mature termite workers are not immediately hindered from ordinary mobility by the commencement of an EPN infection. When immediate immobilization takes place, it is most common with immature workers whose diminutive viscera are more easily disrupted. Once the IJ’s mature into adult males and females, the onset of mechanical injury is inevitable, but for mature termites that does not occur for hours or days, during which time the infected termite worker will most likely have travelled a considerable distance, from the infection site, into other portions of the termite colony’s workings.

Close confines within the micro-workings of the termite superorganism are an artifact of the subterranean termite’s innate thigmophilic nature. The organism prefers to travel and live within a space that allows it to touch all sides of the tubular enclosure at once. Thus a termite worker constantly feels its way along its path, repeatedly palpating the floor, sides, and ceiling of that path with its extremities. This means that traveling workers will come into close, direct contact with whatever occupies its pathway as it passes by, and if that occupant is the cadaver of an EPN-infected termite, the emerging IJ’s have a fighting chance of finding a new host.

EPN host-finding is improved if the termite worker loiters for one reason or another near or in contact with the cadaver of an EPN-infected termite, and is improved again if the viscous fluids that seep from the cadaver, carrying a fresh load of EPN IJ’s, contacts the live worker’s body and sticks to it. Even as the contaminated worker is infected by one or more of the EPN in those fluids, it also distributes them deeper into the colony’s workings, depositing them on others as they pass. If the fluid sticks to the worker at a location near its mouth or anus–the favored ports of entry for Steinernematid EPN–the chance of infection is even better.

Proving Colony Elimination

It is easier to describe how EPN can wipe out termite colonies than it is to prove that they have done it. Nan-yao Su occupied a spot similar to this in the late 1980’s, as his experiments with hexaflumuron began to show success. Dr. Su diligently collected and analyzed field data that eventually proved, to a level of certainty acceptable to most of his colleagues, that hexaflumuron eliminated termite colonies. Our field trials, using EPN-optimized termite interceptors, are producing similar results. As I learned from Su’s experience, however, demonstrating termite colony elimination is a lengthy process, and only patience and diligence will deliver success.

As this field work continued, the design of the TermiteBiotic™ Termite Interceptor, Annunciator, & Inoculator (TIAI), and its method of use, continued to improve.

This device has now been successfully tested throughout Texas, with several species of native subterranean termites as well as the Formosan subterranean termite. In every case, the termite colonies involved have been eliminated through the use of nematode inoculations carried out in our commercial production termite interceptor, annunciator, and inoculator, known as the TermiteBiotic TIAI-IV™.

Our lab studies in Texas have used procedures adapted from those described by Dr. Khuong Nguyen, a biological scientist at the University of Florida entomology and nematology lab, and others. We gratefully acknowledge these contributions. Our studies have specifically tested various configurations of termite interceptors to show that with the TermiteBiotic TIAI-IV™ it is possible to substantially increase the rate of EPN infection, to the point that termite colony elimination, using EPN supplemented with natural mineral and biological adjuvants, is a proven, fully demonstrable, scientific reality.

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References to Relevant Scientific Literature:

• Fodor, A., et al. 1994. Effects of Temperature and Dietary Lipids on Phospholipid Fatty Acids and Membrane Fluidity in Steinernema carpocapsae. Journal of Nematology 26(3):278-285.
• Kaya, Harry K. 2000. Molluscicidal nematodes for biological control of pest slugs. Slosson Report (2000-2001)
• Lacey, L. A., et al. 2001. Insect Pathogens as Biological Control Agents: Do They Have a Future? Biological Control 21:230–248
• Mahar, Ali Nawazi, et al. 2004. Pathogenicity of bacterium, Xenorhabdus nematophila isolated from entomopathogenic nematode (Steinernema carpocapsae) and its secretion against Galleria mellonella larvae. Journal of Zhejiang University Science, 6B(6):457-463.
• Martens, Eric C., et al. 2003. Early Colonization Events in the Mutualistic Association between Steinernema carpocapsae Nematodes and Xenorhabdus nematophila Bacteria. Journal of Bacteriology 185(10):3147-3154.
• Münch, Anna, et al. 2008. Photorhabdus luminescens genes induced upon insect infection. BMC Genomics 2008, 9:229
• Park, Youngjin, and David Stanley. 2006. The entomopathogenic bacterium, Xenorhabdus nematophila, impairs hemocytic immunity by inhibition of eicosanoid biosynthesis in adult crickets, Gryllus Wrmus. Biological Control 38 (2006) 247–253.
• Sicard, Matthieu, et al. 2004. When Mutualists are Pathogens: an Experimental Study of the Symbioses between Steinernema (entomopathogenic nematodes) and Xenorhabdus (bacteria). Journal of Evolutionary Biology 17(5):985-993.
• Sicard, Matthieu, et al. 2006. Interspecific competition between entomopathogenic nematodes (Steinernema) is modified by their bacterial symbionts (Xenorhabdus). BioMed Central Evolutionary Biology 6:68.
• Snyder, Holly, et al. 2007. New Insights into the Colonization and Release Processes of Xenorhabdus nematophila and the Morphology and Ultrastructure of the Bacterial Receptacle of Its Nematode Host, Steinernema carpocapsae. Applied & Environmental Microbiology 73(16):5338-5346.
• Tailliez, Patrick, et al. 2006. New insight into diversity in the genus Xenorhabdus, including the description of ten novel species. International Journal of Systematic and Evolutionary Microbiology 56, 2805–2818.
• Udo-Ehlers, Ralph. 2003. Safety and regulation of entomopathogenic nematodes.
• Vyas, R. V. 2008. Significance of metabolites of native Xenorhabdus, a bacterial symbiont of Steinernema, for suppression of collar rot and root knot diseases of groundnut. Indian Journal of Biotechnology Vol. 7: 171-177.

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