Developing the Arsenal Against Pest and Vector Dipterans: Inputs of Transgenic and Paratransgenic Biotechnologies

Insects are the most numerous of all animals and are found in almost every inhabitable place on earth. The order Diptera (true flies) contains many members that are notorious agricultural pests, nuisance or vectors of diseases. The list is long: mosquitoes, tsetse flies, screw worms, fruit flies, sand flies, blow flies, house flies, gall and biting midges, black flies, leaf miners, horse flies, and so on. Efforts to combat some of these pests and vectors have resulted in control measures such as the chemical, physical, and cultural control methods. These methods, though largely beneficial, have disadvantages and limitations, which sometimes seem to outweigh the problems initially sought to be controlled. The chemical method, for example, is not environment-friendly since it negatively affects many nontarget organisms and disrupts ecosystem balance. Development of insecticide resistance by pests/vectors is another concern. Molecular biotechnology has introduced vast arrays of novel ways to tackle pests and disease vectors, as well as improve the potency of existing control methods. This chapter looks at transgenic and paratransgenic biotechnologies and how they have been applied so far to develop and expand the arsenal against dipteran pests and disease vectors. Further, we discuss the advantages, disadvantages, and limitations of these technologies

Modeling horizontal gene transfer (HGT) in the gut of the Chagas disease vector Rhodnius prolixus

<p>Abstract</p> <p>Background</p> <p>Paratransgenesis is an approach to reducing arthropod vector competence using genetically modified symbionts. When applied to control of Chagas disease, the symbiont bacterium <it>Rhodococcus rhodnii</it>, resident in the gut lumen of the triatomine vector <it>Rhodnius prolixus </it>(Hemiptera: Reduviidae), is transformed to export cecropin A, an insect immune peptide. Cecropin A is active against <it>Trypanosoma cruzi</it>, the causative agent of Chagas disease. While proof of concept has been achieved in laboratory studies, a rigorous and comprehensive risk assessment is required prior to consideration of field release. An important part of this assessment involves estimating probability of transgene horizontal transfer to environmental organisms (HGT). This article presents a two-part risk assessment methodology: a theoretical model predicting HGT in the gut of <it>R. prolixus </it>from the genetically transformed symbiont <it>R. rhodnii </it>to a closely related non-target bacterium, <it>Gordona rubropertinctus</it>, in the absence of selection pressure, and a series of laboratory trials designed to test the model.</p> <p>Results</p> <p>The model predicted an HGT frequency of less than 1.14 × 10^-16per 100,000 generations at the 99% certainty level. The model was iterated twenty times, with the mean of the ten highest outputs evaluated at the 99% certainty level. Laboratory trials indicated no horizontal gene transfer, supporting the conclusions of the model.</p> <p>Conclusions</p> <p>The model treats HGT as a composite event, the probability of which is determined by the joint probability of three independent events: gene transfer through the modalities of transformation, transduction, and conjugation. Genes are represented in matrices and Monte Carlo method and Markov chain analysis are used to simulate and evaluate environmental conditions. The model is intended as a risk assessment instrument and predicts HGT frequency of less than 1.14 × 10^-16per 100,000 generations. With laboratory studies that support the predictions of this model, it may be possible to argue that HGT is a negligible consideration in risk assessment of genetically modified <it>R. rhodnii </it>released for control of Chagas disease.</p

Bacterial symbiosis in arthropods and the control of disease transmission.

Bacterial symbionts may be used as vehicles for expressing foreign genes in arthropods. Expression of selected genes can render an arthropod incapable of transmitting a second microorganism that is pathogenic for humans and is an alternative approach to the control of arthropod-borne diseases. We discuss the rationale for this alternative approach, its potential applications and limitations, and the regulatory concerns that may arise from its use in interrupting disease transmission in humans and animals

Estimate of CRP and TNF-alpha level before and after periodontal therapy in cardiovascular disease patients

Introduction: Epidemiological studies show that individuals with periodontitis have a radically amplified threat to develop cardiovascular disease. CRP&amp; TNF-α, are acute phase proteins monitored as a marker of inflammatory status, which have been identified as a major risk factor for atherosclerotic complications. Elevated CRP &amp; TNF-α level in periodontitis patients have been reported by several groups. The present study was performed to determine whether presence of periodontitis and periodontal therapy could influence the serum levels of CRP &amp; TNF-α in cardiovascular disease patients. Methods: Forty cardiovascular disease subjects participated in the study. They were classified into two groups. Group A (Control) where no periodontal treatment was given, Group B (Test) where periodontal treatment (scaling &amp; root planing) was performed. Periodontal clinical parameters like OHI-S, probing pocket depth, were evaluated together with serum CRP, TNF-α, at baseline and reassessed after 8 weeks for all the subjects in both the groups. Results: The CRP &amp; TNF-α levels in both the groups decreased but the decrease in the Group A was minimal and was not statistically significant (P&gt;0.05); whereas in Group B where periodontal therapy was performed, there was statistically significant decrease. Conclusion: It can be concluded from the study that there can be a possible causal relationship between pathogenesis of periodontal disease and CVD as inferred from the statistical significant outcome in the form of decreased inflammatory biomarkers after the periodontal treatment.Key words: CVD, CRP, TNF-alpha, lipopolysaccharide, periodontiti

Identification of the Midgut Microbiota of An. stephensi and An. maculipennis for Their Application as a Paratransgenic Tool against Malaria

The midgut microbiota associated with Anopheles stephensi and Anopheles maculipennis (Diptera: Culicidae) was investigated for development of a paratransgenesis-based approach to control malaria transmission in Eastern Mediterranean Region (EMR). Here, we present the results of a polymerase chain reaction (PCR) and biochemical-based approaches to identify the female adult and larvae mosquitoe microbiota of these two major malaria vectors, originated from South Eastern and North of Iran. Plating the mosquito midgut contents from lab-reared and field-collected Anopheles spp. was used for microbiota isolation. The Gram-negative and Gram-positive bacterial colonies were identified by Gram staining and specific mediums. Selected colonies were identified by differential biochemical tests and 16S rRNA gene sequence analysis. A number of 10 An. stephensi and 32 An. maculipennis adult mosquitoes and 15 An. stephensi and 7 An. maculipennis larvae were analyzed and 13 sequences of 16S rRNA gene bacterial species were retrieved, that were categorized in 3 classes and 8 families. The majority of the identified bacteria were belonged to the γ-proteobacteria class, including Pseudomonas sp. and Aeromonas sp. and the others were some closely related to those found in other vector mosquitoes, including Pantoea, Acinetobacter, Brevundimonas, Bacillus, Sphingomonas, Lysinibacillus and Rahnella. The 16S rRNA sequences in the current study aligned with the reference strains available in GenBank were used for construction of the phylogenetic tree that revealed the relatedness among the bacteria identified. The presented data strongly encourage further investigations, to verify the potential role of the detected bacteria for the malaria control in Iran and neighboring countries

Cultivation-Independent Methods Reveal Differences among Bacterial Gut Microbiota in Triatomine Vectors of Chagas Disease

Chagas disease is one of the most important endemic diseases of South and Central America. Its causative agent is the protozoan Trypanosoma cruzi, which is transmitted to humans by blood-feeding insects known as triatomine bugs. These vectors mainly belong to Rhodnius, Triatoma and Panstrongylus genera of Reduviidae. The bacterial communities in the guts of these vectors may have important effects on the biology of T. cruzi. For this reason, we analyzed the bacterial diversity hosted in the gut of different species of triatomines using cultivation-independent methods. Among Rhodnius sp., we observed similar bacterial communities from specimens obtained from insectaries or sylvatic conditions. Endosymbionts of the Arsenophonus genus were preferentially associated with insects of the Panstrongylus and Triatoma genera, whereas the bacterial genus Serratia and Candidatus Rohrkolberia were typical of Rhodnius and Dipetalogaster, respectively. The diversity of the microbiota tended to be the largest in the Triatoma genus, with species of both Arsenophonus and Serratia being detected in T. infestans