Instant Insecticide Resistance by Symbiosis

Contributed by: Zhenguo Lin

In the first half of the 20^th century, chemical pesticides began to be widely used to control a variety of pest species. In her 1962 groundbreaking book Silent Spring, Rachel Carson argued that insects are building up resistance to pesticides. Sadly, as Carson foretold, cases of resistance surfaced within two to 20 years after the introduction of every kind of new insecticide.

It is generally believed that the development of pesticide resistance is generated by changes of genetic information in the pest's genomes and it takes many generations for the pesticide resistant genes to spread in the population. However, some short cuts of acquiring pesticide resistance actually exist. As discussed in my previous blog, it is found that European house mice stole a rodenticide resistant gene through hybridization with Algerian mice (1). In a recent PNAS paper (2) , Kikuchi et al. found an unexpected mechanism of rapid acquisition of insecticide resistance from bacterial symbionts in bean bugs and stinky bugs, which are major pests in agriculture.

Fenitrothion is an organophosphate insecticide that acts as an inhibitor of cholinesterase, so that the nerve function is damaged in insects, humans, and many other animals. Fenitrothion has been heavily used to kill a wide range of pests. However, it has been found that repeated application of fenitrothion leads to rapid increase of fenitrothion-degrading microbes, including some species in the bacterial genus of Burkholderia. These bacteria are able to breakdown feritrothion into products that can be used as their carbon source.

Burkholderia are able to inhabit prosperously in the midgut of the bean bugs as symbionts. The infected bean bugs tend to grow bigger than uninfected bugs, showing mutual benefits of symbiosis.  Kikuchi et al. infected the bean bugs with six different Burkholderia species (strains): three of them are fenitrothion-degrading and the others are non-degrading (Figure 1) . They found that the bean bug infected with fenitrothion-degrading Burkholderia has much higher survival rates than those bugs infected with non-degrading Burkholderia, because the degraded fenitrothion is almost non-toxic to bugs. This study indicates that the insects can become resistant to fenitrothion instantly after they swallow these fenitrothion-degrading bacteria.

Interestingly, unlike many other insects, the offspring of bean bugs does not inherit Burkholderia from their mothers. The bean bugs need to pick the Burkholderia symbionts from surrounding soils each generation before reaching the adult stage. This seems to be inefficient, but it reduces the possibility that the fenitrothion-degrading bacteria become so dependent on their hosts that they lose their chemical-detoxifying genes. In addition, the authors found that the resistant Burkholderia species can increase rapidly in soil after treated with fenitrothion even though these bacteria are very rare in natural environments. The fenitrothion-degrading species rapidly become the most dominant group (>80%) in the Burkholderia population after merely one month of fenitrothion treatment. Therefore, bean bugs can easily acquire pesticide resistance because the resistance may have already developed in the bacterial population even before the arrival of insects. In addition, considering that the highly diversified enzymatic functions of bacteria are able to detoxify many different pesticide, the acquisition of instant pesticide resistances from these bacterial symbionts will definitely bring new challenges for the efficiency of insecticides.

Figure 1.

Insecticide resistance of R. pedestris infected with fenitrothion-degrading Burkholderia strains. (A and B) Survival of third instar nymphs of R. pedestris infected with the fenitrothion-degrading and nondegrading Burkholderia strains when reared on fenitrothion-coated soybean seeds. Results under the host genetic background TKS-1 (A) and TKA-7 (B) are shown. Mean and SE of 10 replicates are indicated at each data point. Each asterisk indicates that survival rate of the insects infected with the fenitrothion-degrading Burkholderiastrain is significantly higher than survival rate of the insects infected with the allied nondegrading strain (likelihood ratio test; P < 0.01). (Cand D) Resistance of Burkholderia-infected R. pedestris to percutaneous application (C) and oral administration (D) of fenitrothion. Third instar nymphs, which were infected either with the fenitrothion-degrading Burkholderia strain (SFA1) or with the nondegrading Burkholderia strain (RPE67), were administrated with 30 pmol of fenitrothion, and their survival was inspected 24 h later. On each of the columns is shown number of surviving insects/total number of treated insects. Statistically significant differences in the survival rates are shown (Fisher’s exact probability test). (2)

References

1. Song, Y.,  Endepols, S.,  Klemann, N., Richter, D.,  Matuschka, F.R., Shih, C.H., Nachman, M.W., and Kohn, M.H. 2011. Adaptive introgression of anticoagulant rodent poison resistance by hybridization between Old World mice. Current Biology 21:1296-1301

2. Kikuchi Y, Hayatsu M, Hosokawa T, Nagayama A, Tago K, Fukatsu T. 2012. Symbiont-mediated insecticide resistance. Proc Natl Acad Sci USA 109:8618–8622

3. Kikuchi Y, Hosokawa T, Fukatsu T. 2007. Insect-microbe mutualism without vertical transmission: A stinkbug acquires a beneficial gut symbiont from the environment every generation. Appl Environ Microbiol 73:4308–4316

Rodents with No Y Chromosome and Decay of the Human Y

            In my previous commentary on the sex determination of Tokudaia spiny rats dated May 21, 2012, I mentioned that Amami and Tokunoshima spiny rats (T. osimensis and T. tokunoshimensis) have no Y chromosome and no Sry gene and their sex chromosome type is XO in both males and females and that the gene contents of the male X and the female X chromosomes must be different. However, examining ten genes that are related to sex determination, Kuroiwa et al. (1) concluded that the two chromosomes contain the same set of genes. This suggests that my statement was incorrect (A. Kuroiwa, personal communication).

            However, this does not necessarily exclude the possibility that there may be some sex-determining genes located on the X chromosomes. This possibility is supported to some extent by the mechanism of formation of XO zygotes. Because both males and females have the XO chromosome type, they should produce the gametes with one X or no X chromosome in both sexes. Therefore, the offspring may have genotypes XX, XO, and OO with the probabilities of ¼, ½, and ¼, respectively. However, genotypes XX and OO are not observed, so that they must be lethal. Why is the genotype XX lethal? If the male X (X^m) and the female X (X^f) are identical and normal, the genotype X^mX^f should be viable and be able to establish the XX genotype in the population. One possible explanation for the absence of XX (or X^mX^f) and the persistence of only XO could be that X^m and X^f carry the male and the female-determining genes which are incompatible with each other and therefore X^mX^f individuals will die. Of course, this is an overstretched interpretation at present, and we need to study the genes located on the male X and the female X chromosomes more extensively.

            Kuroiwa et al. identified copies of Cbx2 genes on two autosomal chromosomes in each of the two Tokudaia species, and the number of copies was higher in males than in females. One of the two chromosomes is linked to the Sox9 gene, as in the case of other mammalian species, and therefore the Cbx2 gene located on this chromosome appears to be the original one, and other Cbx2 genes located on the other chromosome are considered to be the genes transferred from the original chromosome and then duplicated. Kuroiwa et al. speculated that these duplicate Cbx2 genes are responsible for testis formation because the Cbx2 genes are known to suppress ovary development in humans and mice. Kuroiwa (personal communication) then suggests that the new chromosome carrying more Cbx2 genes has become a neo-Y chromosome and the homologous chromosome carrying a smaller number of Cbx2 genes has become a neo-X chromosome.  

            This is an interesting suggestion, but it is not without deficiencies. First, there is no proof that a larger number of Cbx2 copies is really responsible for sex determination. Second, there must be a recombination reduction in the duplicate gene region to keep the duplicate genes together. Otherwise, the neo-Y chromosome cannot be isolated from the neo-X chromosome (2). Once no recombination system evolves in this region, lethal mutations are expected to accumulate particularly in small populations (3). At the present time, no such evidence seems to exist.

            Of course, identification of the sex determination gene in these species would be very difficult because the population size of Amami and Tokunoshima spiny rats is very small and protected by the Japanese government. Therefore, one cannot get even small samples easily. Nevertheless, it would be important to clarify the evolutionary mechanism of the XO/XO type, because this would give some insight into the general pattern of Y chromosome evolution.

Fig.1. Degeneration of the sex-specific element (Y or W) from an original autosome (purple) to form more or less differentiated XY (blue) or ZW (pink) sex chromosome pairs. Degraded Y chromosomes harbor male-specific genes (blue lines) and W harbor female-specific genes (pink lines). Examples of vertebrate species that exhibit this level of differentiation of XY or ZW chromosomes are given on the left or right, respectively. Dotted lines represent pairing and recombination.      From Graves (5).

            Jennifer Graves (4; 5; 6) has repeatedly argued that the human Y chromosome is subject to a high rate of deleterious mutation and appears to become extinct in about ten million years. It is known that only 50 out of the 1,500 genes that existed in the original Y chromosome are surviving currently in the human chromosome. She states: “Accelerated degeneration of the Y chromosome is found in 5 – 15% of severely infertile men whose infertility is caused by wholesale deletion of parts of this chromosome.” She envisages that once the Y chromosome is lost, a new set of sex chromosomes like those of Tokudaia spiny rats must be generated.

            In principle, I disagree with her, because strong purifying is operating for keeping all Y chromosome genes required for male fertility and formation of male phenotype. However, if contraception practice becomes more popular and each married couple produces a smaller number of offspring, the purifying selection will be weaker. It is then possible that infertile men increase and the probability of decay of the Y chromosome genes may be enhanced. In this case the formation of a new sex determination system similar to that of Tokudaia could be one solution. However, because humans can control their evolution, it is unlikely for this event to happen.

References

1. Kuroiwa A, Handa S, Nishiyama C, Chiba E, Yamada F, Abe S, and Matsuda Y. 2011. Additional copies of CBX2 in the genomes of males of mammals lacking SRY,the Amami spiny rat (Tokudaia osimensis) and the Tokunoshima spiny rat (Tokudaia tokunoshimensis). Chromosome Res 19:635-644.

2. Nei, M. 1969. Linkage modification and sex difference in recombination. Genetics 63:681-699.

3. Nei, M. 1970. Accumulation of nonfunctional genes on sheltered chromosomes. Am. Nat. 104:311-322.

4. Aitken RJ, and Marshall Graves JA. 2002. The future of sex. Nature 415:963.

5. Graves JAM. 2008. Weird animal genomes and the evolution of vertebrate sex and sex chromosomes. Ann Rev Genet 42:565-586.

6. Griffin DK. 2012. Is the Y chromosome disappearing?--both sides of the argument. Chromosome Res 20:35-45.