Pesticides are substances widely used in the field of agriculture and health for the elimination of food pests, reduction of food losses, and control of vectors transmitting human and animal diseases ^[1][2][1,2]. Currently, due to the increase in the world population and its need to produce more food, as well as to avoid losses in agricultural crops, the use of pesticides has increased. Organophosphates pesticides (OPs) belong to the group of the most widely used pesticides, which are derived from phosphorous compounds, such as phosphoric and phosphorothioic acid. Globally, about 2 million tons of pesticides are used every year; however, as of 2020, the annual increase of these substances was estimated at 3.5 million tons, of which approximately 40% represent OPs ^[1][3][1,3]. Additionally, an estimate of more than 3 million people are exposed to OPs each year, leading to 300,000 deaths worldwide [4].

Although OPs have been used to control pests for more than 50 years [5], the use of OPs has increased considerably since the banning of organochlorine pesticides, because OPs have limited persistence in the environment and pose a lower risk to human health compared to organochlorines; however, excessive use, poor storage, transport, application, and disposal of residues pose a latent risk of affecting non-target organisms ^[6][7][8][6,7,8]. OPs are incorporated into organisms by three routes of exposure: oral, respiratory, and dermal. In terms of absorption, the main absorption route is through the diet, while respiratory absorption will depend on physicochemical properties and environmental persistence. On the other hand, dermal absorption is influenced by polarity and solubility. Once absorbed, the compound is distributed in the organism, and subsequently bio-transformed to more hydrophilic metabolites to increase polarity and facilitate elimination. The main organ where biotransformation takes place is the liver, although this phenomenon also occurs in the kidney, intestine, and gills ^[9][10][9,10]. In vertebrates, biotransformation is a process that takes place in different stages, involving both bioactivation and detoxification [11]. OPs are mainly bio-transformed in the liver via CYP450 (phase I) by chemical reactions (oxidative desulfurization), to form highly toxic compounds, such as oxon metabolites. The metabolic process of detoxification consists of reactions (phase II) of dearylation and hydrolysis to convert the oxon into dialkyl phosphates (DAPs), dialkyl thiophosphates (DATPs), and 2-isopropyl-4-methyl-6-hydroxy pyrimidine (IMPH), which are highly soluble metabolites that can be excreted mainly through urinary conjugation reactions ^{[7][8][12][13]}[7,8,12,13].

2. Immune Response

Life began on our planet more than 3.5 billion years ago, and evolving single-cell organisms, archaea, bacteria, and eukaryotes, have flourished ever since. Around 600 million years ago, multicellular organisms (metazoans) began to form in conjunction with a dramatic increase in atmospheric oxygen levels. This development was followed by a remarkable diversification of metazoan species in such a relatively short period that has been called the “evolutionary big bang.” ^[14][33]. The evolutionary emergence of vertebrates was accompanied by major morphological and functional innovations, including the development of an immune system ^[15][34]. Innate immunity is the first line of defense against antigens ^[16][35]. Innate mechanisms are the first to respond to an antigen, recognizing it in a generic and non-specific way, thus generating a rapid response to eliminate it, but it does not provide long-term protection ^[17][18][19][36,37,38]. Innate immunity includes effector molecules, such as interferons, complement proteins, natural antibodies, growth inhibitors, and protease inhibitors, and cells such as macrophages, monocytes, neutrophils, and mast cells. Additionally, included are species-specific physical barriers such as mucus, skin, gills, intestines, and nostrils ^[18][37]; whereas, the cellular defense of invertebrates is carried out by hemocytes through phagocytosis, cytotoxic reactions that include the release of lysosomal enzymes and antimicrobial peptides, and respiratory burst ^[20][39]. Innate immunity is considered evolutionarily older than adaptive immunity ^[16][35]. Even unicellular organisms have heritable defense mechanisms, and every multicellular organism appears to have a complex innate immune system ^[21][40]. The basic protective strategy of an innate immune system is for the organism to constitutively produce generic receptors that recognize conserved patterns on different classes of pathogens to trigger an inflammatory response that limits pathogen invasion ^[14][33]. Innate immunity is of paramount importance for both invertebrates and some lower vertebrates, such as fish ^[19][38], to the extent that it has been suggested that teleost fish have a more robust innate response than mammals ^[16][35]. In addition, they have an instructive role for adaptive immunity mechanisms ^[19][38]. Adaptive immunity is composed of highly specialized cells, which include populations of lymphocytes, such as B cells, T cells, and natural killer (NK) cells, as well as immunoglobulins and the major histocompatibility complex (MHC) system ^[16][17][18][35,36,37]. Lymphocytes, the specialized cell type of the adaptive immune system, use their cell-surface receptors to recognize antigenic configurations of specific pathogens and then respond to the antigen triggering by clonal amplification, cellular differentiation, and antibody production with the same antigen-binding specificity ^[14][33]. The TCRs recognize peptide fragments of antigens presented by other cells within cell-surface molecules encoded by the major histocompatibility complex (MHC) class I and class II genes. T cells, therefore, typically recognize antigens that have been partially digested by the antigen-presenting cells, primarily dendritic cells, phagocytic cells, and B cells. The membrane-bound and secreted antibodies made by B lineage cells, by contrast, recognize exposed determinants (epitopes) of intact molecules, including surface protein and carbohydrate moieties of invasive microbes ^[14][33]. Even cartilaginous fish, such as sharks, have TCR and BCR genes divided into V, D, J, and constant I region segments and RAG1/RAG2, MHC I, and MHC II genes ^[14][33]. A key feature of adaptive immunity is the development of immunological memory, as memory cells are generated that provide long-lasting specific immunity, and thus play a crucial role in protecting against recurrent infections with a rapid, intense, and efficient response ^[17][18][19][36,37,38], leading to an appropriate response in subsequent encounters with the antigen ^[22][41]. Interestingly, memory formation, previously thought to be a defining feature of adaptive immunity, also occurs in the context of innate immune responses and can be observed even in unicellular organisms, demonstrating the convergent evolutionary history of different aspects of adaptive immunity ^[15][34].

3. OP-Mediated Modulation of the Immune Response

3.1. OP-Mediated Modulation of the Innate Immune Response in Invertebrates

OPs are also able to decrease innate functions of hemocytes in invertebrates. Notwithstanding the above, OPs dysregulate immune functions, and in line with this, the hemocyte count is altered in Drosophila melanogaster Meigen exposed to acephate (acute and chronic) ^[23][24][25][42,43,44]. Contrastingly, monocrotophos, dimethoate, and methyl parathion increased the total hemocyte count in Rhynocoris kumarii Ambrose and Livingstone ^[26][45]. Moreover, hemocyte count alterations have been reported in shrimps (Litopenaeus vannamei Boone) exposed to malathion ^[27][46]. In this regard, azamethiphos and dimethoate decreased the phagocytic index in the marine mollusk Mytilus edulis L. ^[28][29][49,50]. Alongside this, in gastropods Planorbarius corneus L. and Biomphalaria glabrata Say, exposed to chlorpyrifos, the hemocyte count and lysozyme activity were not significantly altered; notwithstanding, functional parameters (phagocytosis and ROS production) were altered ^[20][39]. Futher, hemocyte viability and phagocytic activity decreased after azinphos-methyl exposure in the freshwater snail Chilina gibbosa G. B. Sowerby I. ^[30][51]. In the mussel Diplodon chilensis Gray, the exposure to azinphos-methyl modified the hemocyte count, enzymatic activity (lysozyme and phenoloxidase, and glutathione S-transferase), and phagocytic activity ^[31][52].

3.2. OP-Mediated Modulation of the Innate Immune Response of Vertebrates

To date, information on the immuno-toxic effect of OPs on agnathans and Chondrichthyes is scarce. Fish have been used as indicator species for water quality contamination, because fish are organisms closely related to the aquatic environment and suffer stress and disturbance due to the presence of pollutants ^[32][61]. Therefore, organisms such as medaka, zebrafish, carp, guppy, and tilapia are often used as models for toxicological testing ^[33][62]. Studies have focused on the toxic effect on bony fish. OPs induce changes in fish immune cell counts, as has been observed in invertebrates. In this regard, white blood cell (WBC) counts increased in common carp (Cyprinus carpio carpio L.) exposed to chlorpyrifos ^[34][63]. Alongside that, other immune cell counts can be modulated, as reported in Oncorhynchus mykiss Walbaum exposed to diazinon, where monocytes were lower, while a significant increase in neutrophils was observed ^[35][64]. In line with this, neutrophilia induced by diazinon was also described in Pangasius hypophthalmus Sauvage ^[36][65]. Hematological parameters (WBCs and heterophils), respiratory burst, lysozyme activity, and C-reactive protein were also affected in O. niloticus exposed to diazinon ^[37][66]. Other alterations to immune mechanisms by OPs exposure are phagocytic parameters. In line with this, malathion has a negative effect on the innate immune response (WBC, ROS production, and phagocytic activity) in C. carpio carpio ^[38][75]. Additionally, acute and chronic exposure to chlorpyrifos effects leukocyte phagocytic capacity ^[39][40][41][76,77,78]. Furthermore, malathion reduces the phagocytic capacity of Murray codfish (Maccullochella peelii T. L. Mitchell) ^[42][79]. In addition, phagocytosis was reduced, and an ROS production increment was reported in O. niloticus exposed to diazinon ^[43][28]. Immunosuppressive effects, such as an enzyme activity levels decrement, have been demonstrated in populations of Leptodactylus latrans and Hyloxalus pulchellus ^[44][80]. In terms of cell counts, a drastic reduction in WBC content was observed in Bufo melanostictus Schneider (Common Indian toad) exposed to malathion ^[45][81]. In Rana pipiens Schreber, a significant decrease in splenocyte numbers (cellularity), and phagocytic activity was observed in frogs sampled at pesticide-impacted sites ^[46][82]. In Rana temporaria, a lower number of blood leukocytes was also observed when they were exposed to high concentrations of tetrachlorvinphos compared to unexposed animals ^[47][83]. Studies on the effect of OPs on the mechanisms of innate immunity in reptiles are scarce; the reports published to date are presented below. On glyphosate-based formulation exposition of Caiman latirostris, a decrease in WBC counts, as well as a higher percentage of heterophils, was reported ^[48][84]. In the Caspian pond turtle (Mauremys caspica caspica Gmelin) exposed to diazinon, a reduction in serum complement, lysozyme activity, and phagocytosis were reported, while the heterophil/lymphocyte ratio increased ^[49][85]. Moreover, in broad-snouted caiman (Caiman latirostris Daudin), a lower complement system activity was reported after commercial-mixed glyphosate exposure ^[50][86]. A multiple exposure assay of glyphosate, chlorpyrifos, and cypermethrin, during embryo development of tegu lizard (Salvator merianae Duméril and Bibron) detected a decrease in heterophils and the heterophil/lymphocyte ratio, while natural antibody titers increased ^[51][87]. Birds are non-target species and OPs exert immunotoxicity. Methidathion and chlorpyrifos exposure in young chickens reduces WBC and neutrophil count ^[52][88]. Moreover, in broiler chicks, exposure to monocrotophos reduces active splenic macrophages ^[53][89]. Chlorpyrifos administered to broiler chicks caused a dose-dependent decrease in phagocytic activity ^[54][90]; while no difference in the innate immune response in sub-chronic exposure to malathion of birds (Coturnix coturnix japonica Temminck and Schlegel) was reported ^[55][91].

3.3. OP-Mediated Modulation of the Adaptive Immune Response in Vertebrates

The parameters of adaptive immunity and their alteration in fish due to exposure to OPs have been little studied. In this regard, ^[56][106] reported that in vitro exposure to diazinon caused a decrease in plasma IgM in Nile tilapia (O. niloticus). Similarly, in carp (C. carpio) exposed to glyphosate-POEA, IgM mRNA levels were lower than in the control group ^[57][107]. 

Given that amphibians are frequently exposed to agricultural pesticides, it is possible that these pollutants alter their immune system and render them more susceptible to different pathogens ^[58][60]. In the specific case of OPs, exposure to chlorpyrifos in toad tadpoles (Odontophrynus carvalhoi) altered the WBC, which could alter the ability of tadpoles to respond to environmental stress, make them more susceptible to infection by various pathogens, and thus reduce their chances of survival ^[59][109].

It has been described that pesticides may cause negative effects and alterations in non-target species. In particular, the effect of quinalphos on humoral immune response in chickens was tested. In theis study, all chicks were vaccinated with the Ranikhet disease vaccine on day 4 and Infectious bursal disease (IBD) on day 15. In addition, a group of chicks was given quinalphos in feed. There was a significant suppression in serum globulin, gamma globulin, and specific antibody titre against Ranikhet disease and IBD. B-lymphocyte blastogenesis was found to be reduced in chicks fed quinalphos in comparison to controls, indicating an immunosuppressive effect ^[60][111].

Exposure to OPs has been described to deregulate both B-cell maturation and T-cell differentiation in mammals. In this regard, IgM concentration, cytotoxic T lymphocyte count, IFNγ, and TNF-α production were decreased in mice exposed to parathion. In addition, Th2 cytokine production and GATA-3 gene expression were significantly increased ^[61][117]. Alongside that, increased expression of surface T-cell receptors and levels of Th1 cytokines (IFN-γ, TNF-α) and exacerbation of T-lymphocyte-mediated allergic reactions were induced in rats ^[62][118]. In rats exposed to fenitrothion, serum levels of TNF-α and IL-2 were increased, while it caused a reduction of IgG and IgM ^[63][119].

4. Conclusions

The immuno-toxic effects mediated by OPs affect both invertebrate and vertebrate organisms, with direct repercussions on the innate and adaptive immunity of exposed organisms. Acute exposure has been reported to cause impairments in phagocytosis, respiratory burst, ROS release, hemocyte/leukocyte cellular response, antibody production, cell proliferation, and cytokine release. Hence, exposure to OPs can cause alterations in the various cells of the immune system, which can result in increased susceptibility to infections caused by opportunistic microorganisms, including viruses, bacteria, parasites, and fungi, thus necessitating studies in which the exposure to OPs in relation to an antigenic challenge is evaluated, causing an imbalance in the environment and in the health of organisms.