**5. Pharmacology and therapeutic uses of venom form ants**

**3. Clinical aspects of ants' stings**

Applications

210

Many insect stings are associated with local pathophysiological events, characterized by pain, swelling and redness at the sting site for about 1-2 days [18]. The most severe reactions are associated with allergic disorders, presenting neutrophilic and eosinophilic infiltration and specific IgE production [19]. These manifestations are common in accidents with Hymenoptera insects. Most studies that describe the clinical aspects of ant stings reported accidents with ants of the genus *Solenopsis* (Myrmicinae), known as fire ants [20,21,22]. In most serious cases, these accidental encounter with fire ants can promote multiple body rash, seizures, heart failure,

An Integrated View of the Molecular Recognition and Toxinology - From Analytical Procedures to Biomedical

Accidents with ants of the Ponerinae subfamily are rare or rarely reported. In fact, several concomitant or sequential stings are necessary in order to produce significant clinical symp‐ toms of envenomation, in giant ants, multiple attacks are less probable, since workers have a solitary foraging behavior. However, some of the accidents with giants ants may have medical importance, such as the ones produced by the genus *Paraponera* and *Dinoponera*, popularly known as "true tocandira" and "false tocandira", respectively. Their stings are extremely painful and can cause potentially systemic manifestations such as fever, cold sweats, nausea, vomiting, lymphadenopathy and cardiac arrhythmias [8,25,26]. According to [27,28] the

The ant's venoms have been investigated in a relatively small number of species. In the group of stinging ants, the most investigated species belong to the Myrmeciinae, Ponerinae, Pseu‐ domyrmecinae and Myrmicinae subfamilies. They produce aqueous solutions of proteina‐ ceous venoms containing enzymatic and non-enzymatic proteins, free amino-acids and small biologically active compounds like histamine, 5-hydroxytryptamine, acetylcholine, norepi‐ nephrine, and dopamine [16,17]. Venoms with proteinaceous components are considered as most primitive and are consequently found in other aculeate hymenopterans like wasps and bees [4,16]. A notable exception to this proteinaceous nature of the venom in ants with sting is found in ants of the genera *Solenopsis* (fire ants) and *Monomorium* (Myrmicinae) that produce alkaloid-rich venoms with few proteins. In the Formicinae ants (ex: *Camponotus*, *Formica*), the sting is no more presented, but the poison gland produces a mixtures of simple organic acids an aqueous solution. Formic acid is presented in concentrations up to 65% along with some

As a member of a group of predatory ants (Ponerinae), it is expected that *Dinoponera* would produces such a kind of proteinaceous venom. However, until now few studies have been done with *Dinoponera* venoms. In two of these studies, which compared venoms of a variety of hymenopterans, the presence of proteins, some with enzyme activities (phospholipase A, hyaluronidase, and lipase), was shown for *D. grandis* (in fact, *D. gigantea*) venom [16,29]. In a more recent study, in which the peptide components from the venom of *D. australis* was

and serum sickness nephritis and, more rarely, acute renal failure [23,24].

venom of these ants may be neurotoxic for other insects.

peptides and free amino-acids [16,17].

**4. Venom composition and pharmacological properties**

The first reported case about the therapeutic use of venoms from ants were to treat rheumatoid arthritis. In fact, insects might have components that justify its use in traditional medicine in countries of East Asia, Africa and South America [36]. Lately, several studies of ant venom aimed to demonstrate their beneficial intrinsic properties such as reduction of inflammation, pain relief, improved function of the immune system and liver [37,38].

As the venom from Ponerinae subfamily is composed of a complex mixtures of proteins and neurotoxins [39] we would expected to have several pharmacological properties. Small peptides isolated from *Paraponera clavata* venom, called poneratoxin (PoTx) interfere with sodium channels function and have potential use as a biological insecticide [40,41].

Several distinctive pharmacological activities were demonstrated with peptides isolated from *Pachycondyla goeldii* and *Myrmecia* sp. In one of these works, antimicrobial activity against both Gram positive and Gram negative bacteria was observed [42, 43]. In a recent study [44], it was reported that the venom from *Pachycondila sennaarensis* has a significant antitumor effect on breast cancer cells in a dose and time dependent manner without affecting the viability of non tumor cells. In addition, some studies have also shown the renal effects of Hymenoptera venoms. In fact, in more serious accidents with venoms from wasps and bees acute renal failure generally occurs [45,46, 47, 48].

### **6. Genomic study of ant venom composition**

Since the description of DNA double helix by Francis Crick and James Watson (1953), re‐ combinant DNA technology and genomics revolutionized numerous areas of life science. The comprehension of the biochemical and molecular basis of inheritance had been improved our knowledge about the complexity of all forms of life and the manner how genes and proteins interact to create diversity. The genomic revolution was additionally expanded with the advent of bioinformatic, the 'omic' science (transcriptomic, proteomic, peptidome, metabolomic, glycome) and, presently, system biology.

Collective efforts have been joined to annotate the gene composition of insects. The first complete sequenced genome of insect was from the fruit fly *Drosophila melanogaster*, in 2000, followed by a flurry of activities aimed at sequencing the genomes of several additional insect species. In the field of toxinology, the hymenopterans are receiving special attention due to their behavior and the ability to produce venom.

**DQv**

Molecular Pharmacology and Toxinology of Venom from Ants

http://dx.doi.org/10.5772/53539

213

 **50**

**37**

**25**

 **20**

 **15**

trophoresis visualized with Comassie Brilliant Blue.

and a report will be published elsewhere.

mechanisms related to the observed effects.

vDq \*

**%pTNa+**

**0.4** Ctrl

**<sup>30</sup> <sup>60</sup> <sup>90</sup> <sup>120</sup> 0.0**

**Time (min)**

**0.1 0.2 0.3**

**UF (mL.g-1.min-1)**

**Figure 3.** Electrophoretic profile of *Dinoponera quadriceps* total venom (DQv) in one-dimensional SDS-PAGE gel elec‐

The peptide mass fingerprint (PMF), as well as other proteomic analysis is being conducted

Pharmacological studies have been realized with *Dinoponera quadriceps* venom, particularly, in a system of isolated perfused rat kidney. We now know that at concentrations of approxi‐ mately 10μg/mL increased urinary flow, glomerular filtration rate and decreased vascular resistance and sodium tubular transport, suggesting a natriuretic and diuretic effect. Further‐ more, in studies with renal tubule cells (MDCK - Madin-Darbin Canine Kidney) the same venom induced cell cytotoxicity, on MTT assay (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenylte‐ trazolium bromide) at a dose and time dependent manner. Interestingly, greater cytotoxicity was observed in the shorter incubation periods, suggesting that the cell culture could recover after a given exposure time. Additional assays have been designed to evaluate the biological and pharmacological activity of purified component of this venom, as well as highlighting the

**<sup>30</sup> <sup>60</sup> <sup>90</sup> <sup>120</sup> <sup>0</sup>**

**Time (min)**

**Figure 4.** Effect of *D. quadriceps* total venom (DQv) on Urinary flow (UF; A), sodium tubular transport percent(%pTNa; B) and renal vascular resistence (RVR; C). Ctrl=control. Results are expressed as means ± S.E.M., \*p<0.05 (ANOVA).

**<sup>100</sup>** Crtl

vDq \*

DQv DQv DQv

**2 4 6**

**RVR (mmHg/mL .g-1. min-1)**

**<sup>30</sup> <sup>60</sup> <sup>90</sup> <sup>120</sup> <sup>0</sup>**

**Time (min)**

**<sup>8</sup>** Crtl

\* \*

vDq

Up to now, at least 10 ant species had their genomes analyzed and published. The ants whose genomes were sequenced include: the fire ant *Solenopsis invicta* found in South America, United States, China, Taiwan, Australia [49]; the Argentine ant *Linepithema humile*[50], the leaf-cutting ant *Acromyrmex echinator* [51] and *Atta cephalote* [52] found in South America; the red harvester *Pogonomyrmex barbatus* found in North and South America [53], the florida carpenter ant *Camponotus floriandus* from United States; and, the jumper ant *Harpegnatos saltator* from India, Sri Lanka and Southeast Asia [54]. Those ant genomes have provided hundreds of new available nucleotide data.

Apart of a detailed genome analysis, the construction of cDNA libraries from ants' venom glands is an important tool in order to analyze venom composition and discover new molecules that could have biological and pharmacological properties. But an important question arises: why hymenopteran venoms? As we pointed at the beginning of this chapter, there are several reports that hymenopteran venom could have biological properties useful for medical purpos‐ es. In this scope, from traditional and modern medicine reports, description can be found not only about clinical manifestation caused by hymenopterans venom, as allergic response, but al‐ so the benefits of ant venom to treat disease like rheumatoid arthritis and pain [36].

Genomic and transcriptomic studies of hymenopteran cDNA libraries would provide useful information about their protein constituents. Some of these informations would include signal peptide sequences and the presence of post-translational modifications, which cannot be predicted by the studies of mature proteins. Ants genomic studies have shown a number of substances involved in the biology of these insects, such as: vittelogenins, gustatory and odorant receptors, molecules involved in immune response, as well as metabolic and structural proteins like cytochrome P450.
