Interruption of vector transmission by native vectors and “the art of the possible”

In a recent article in the Reader’s Opinion, advantages and disadvantages of the certification processes of interrupted Chagas disease transmission (American trypanosomiasis) by native vector were discussed. Such concept, accepted by those authors for the case of endemic situations with introduced vectors, has been built on a long and laborious process by endemic countries and Subregional Initiatives for Prevention, Control and Treatment of Chagas, with Technical Secretariat of the Pan American Health Organization/World Health Organization, to create a horizon target and goal to concentrate priorities and resource allocation and actions. With varying degrees of sucess, which are not replaceable for a certificate of good practice, has allowed during 23 years to safeguard the effective control of transmission of Trypanosoma cruzi not to hundreds of thousands, but millions of people at risk conditions, truly “the art of the possible.”

The Plasmodium bottleneck: malaria parasite losses in the mosquito vector

Nearly one million people are killed every year by the malaria parasite Plasmodium. Although the disease-causing forms of the parasite exist only in the human blood, mosquitoes of the genus Anopheles are the obligate vector for transmission. Here, we review the parasite life cycle in the vector and highlight the human and mosquito contributions that limit malaria parasite development in the mosquito host. We address parasite killing in its mosquito host and bottlenecks in parasite numbers that might guide intervention strategies to prevent transmission.

Intrusive versus domiciliated triatomines and the challenge of adapting vector control practices against Chagas disease

Chagas disease prevention remains mostly based on triatomine vector control to reduce or eliminate house infestation with these bugs. The level of adaptation of triatomines to human housing is a key part of vector competence and needs to be precisely evaluated to allow for the design of effective vector control strategies. In this review, we examine how the domiciliation/intrusion level of different triatomine species/populations has been defined and measured and discuss how these concepts may be improved for a better understanding of their ecology and evolution, as well as for the design of more effective control strategies against a large variety of triatomine species. We suggest that a major limitation of current criteria for classifying triatomines into sylvatic, intrusive, domiciliary and domestic species is that these are essentially qualitative and do not rely on quantitative variables measuring population sustainability and fitness in their different habitats. However, such assessments may be derived from further analysis and modelling of field data. Such approaches can shed new light on the domiciliation process of triatomines and may represent a key tool for decision-making and the design of vector control interventions.

Atlas of Mexican Triatominae (Reduviidae: Hemiptera) and vector transmission of Chagas disease

Chagas disease is one of the most important yet neglected parasitic diseases in Mexico and is transmitted by Triatominae. Nineteen of the 31 Mexican triatomine species have been consistently found to invade human houses and all have been found to be naturally infected with Trypanosoma cruzi. The present paper aims to produce a state-of-knowledge atlas of Mexican triatomines and analyse their geographic associations with T. cruzi, human demographics and landscape modification. Ecological niche models (ENMs) were constructed for the 19 species with more than 10 records in North America, as well as for T. cruzi. The 2010 Mexican national census and the 2007 National Forestry Inventory were used to analyse overlap patterns with ENMs. Niche breadth was greatest in species from the semiarid Nearctic Region, whereas species richness was associated with topographic heterogeneity in the Neotropical Region, particularly along the Pacific Coast. Three species,Triatoma longipennis, Triatoma mexicana and Triatoma barberi, overlapped with the greatest numbers of human communities, but these communities had the lowest rural/urban population ratios. Triatomine vectors have urbanised in most regions, demonstrating a high tolerance to human-modified habitats and broadened historical ranges, exposing more than 88% of the Mexican population and leaving few areas in Mexico without the potential for T. cruzitransmission.

Assessing the mitochondrial DNA diversity of the Chagas disease vector Triatoma sordida (Hemiptera: Reduviidae)

Triatoma sordida is a species that transmits Trypanosoma cruzi to humans. In Brazil, T. sordida currently deserves special attention because of its wide distribution, tendency to invade domestic environments and vectorial competence. For the planning and execution of control protocols to be effective against Triatominae, they must consider its population structure. In this context, this study aimed to characterise the genetic variability of T. sordida populations collected in areas with persistent infestations from Minas Gerais, Brazil. Levels of genetic variation and population structure were determined in peridomestic T. sordida by sequencing a polymorphic region of the mitochondrial cytochrome b gene. Low nucleotide and haplotype diversity were observed for all 14 sampled areas; π values ranged from 0.002-0.006. Most obtained haplotypes occurred at low frequencies, and some were exclusive to only one of the studied populations. Interpopulation genetic diversity analysis revealed strong genetic structuring. Furthermore, the genetic variability of Brazilian populations is small compared to that of Argentinean and Bolivian specimens. The possible factors related to the reduced genetic variability and strong genetic structuring obtained for studied populations are discussed in this paper.

New record of Pterotaenia fasciata (Wiedemann) (Diptera, Ulidiidae) in Brazil, a probably mechanical vector of enteric bacteria

Pterotaenia fasciata is commonly recorded in rural areas in Argentina, but during a Diptera survey study developed in a reservoir which retains storm water from polluted canals in an urban area of Taboão da Serra municipality, SP, Brazil, we could capture P. fasciata adults. Enteric bacteria Escherichia coli T. Escherich, 1885 and Proteus sp. were isolated from P. fasciata collected in traps inside the reservoir and around it. Fecal coliforms and E. coli were found in the water of the reservoir. These records suggest that a high abundance of this species at urban areas with inadequate sewage canals should reveal these muscoid dipterans as mechanical vectors of enteric bacteria.

Vector competence of Culex quinquefasciatus Say, 1823 exposed to different densities of microfilariae of Dirofilaria immitis (Leidy, 1856)

The metropolitan region of Recife, Brazil is endemic for Dirofilaria immitis and has an environment favorable to the development of Culex quinquefasciatus. The goal of this study was to evaluate the vector competence of the Cx. quinquefasciatus RECIFE population for D. immitis transmission. A total of 2,104 females of Cx. quinquefasciatus RECIFE population were exposed to different densities of D. immitis microfilariae blood meals, ranging from 1,820 to 2,900 mf/ml of blood, in a natural membrane apparatus. The results showed a variation between 92.3% and 98.8% of females fed. The exposure of the Cx. quinquefasciatus RECIFE population to different densities of microfilariae did not influence the mortality of the mosquitoes. Infective larvae from D. immitis were observed in the Malpighian tubules beginning on the 12th day, whereas larvae were observed in the head and proboscis beginning on the 13th day following infection. The vector efficiency index (VEI) presented by the mosquitoes ranged from 7.8 to 56.5. The data demonstrates that the Cx. quinquefasciatus RECIFE population has great potential for the transmission of D. immitis, as it allowed the development of the filarid until the infectious stage at the different densities of microfilariae to which it was exposed.

Spatial evaluation of larvae of Culicidae (Diptera) from different breeding sites: application of a geospatial method and implications for vector control

Spatial evaluation of Culicidae (Diptera) larvae from different breeding sites: application of a geospatial method and implications for vector control. This study investigates the spatial distribution of urban Culicidae and informs entomological monitoring of species that use artificial containers as larval habitats. Collections of mosquito larvae were conducted in the São Paulo State municipality of Santa Bárbara d' Oeste between 2004 and 2006 during house-to-house visits. A total of 1,891 samples and nine different species were sampled. Species distribution was assessed using the kriging statistical method by extrapolating municipal administrative divisions. The sampling method followed the norms of the municipal health services of the Ministry of Health and can thus be adopted by public health authorities in disease control and delimitation of risk areas. Moreover, this type of survey and analysis can be employed for entomological surveillance of urban vectors that use artificial containers as larval habitat.

Rotenoids from Tephrosia toxicaria with larvicidal activity against Aedes aegypti, the main vector of dengue fever

In the search for new larvicides from plants, we have investigated the potential activity of the rotenoids deguelin (1), 12a-hydroxy-α-toxicarol (2) and tephrosin (3), isolated from the bioactive ethanol extract of roots of Tephrosia toxicaria Pers., against Aedes aegypti, the main vector of dengue. The absolute configuration of these compounds was determined by circular dichroism (CD) spectra. The LC50 values of the compounds evaluated justify the potential of T. toxicaria as a new natural larvicide.

PREDICTING THE BOILING POINT OF PCDD/Fs BY THE QSPR METHOD BASED ON THE MOLECULAR DISTANCE-EDGE VECTOR INDEX

The quantitative structure property relationship (QSPR) for the boiling point (Tb) of polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans (PCDD/Fs) was investigated. The molecular distance-edge vector (MDEV) index was used as the structural descriptor. The quantitative relationship between the MDEV index and Tb was modeled by using multivariate linear regression (MLR) and artificial neural network (ANN), respectively. Leave-one-out cross validation and external validation were carried out to assess the prediction performance of the models developed. For the MLR method, the prediction root mean square relative error (RMSRE) of leave-one-out cross validation and external validation was 1.77 and 1.23, respectively. For the ANN method, the prediction RMSRE of leave-one-out cross validation and external validation was 1.65 and 1.16, respectively. A quantitative relationship between the MDEV index and Tb of PCDD/Fs was demonstrated. Both MLR and ANN are practicable for modeling this relationship. The MLR model and ANN model developed can be used to predict the Tb of PCDD/Fs. Thus, the Tb of each PCDD/F was predicted by the developed models.

The yellow fever 17D vaccine virus: molecular basis of viral attenuation and its use as an expression vector

The yellow fever (YF) virus is the prototype flavivirus. The use of molecular techniques has unraveled the basic mechanisms of viral genome structure and expression. Recent trends in flavivirus research include the use of infectious clone technology with which it is possible to recover virus from cloned cDNA. Using this technique, mutations can be introduced at any point of the viral genome and their resulting effect on virus phenotype can be assessed. This approach has opened new possibilities to study several biological viral features with special emphasis on the issue of virulence/attenuation of the YF virus. The feasibility of using YF virus 17D vaccine strain, for which infectious cDNA is available, as a vector for the expression of heterologous antigens is reviewed

The challenge of vector development in gene therapy

Gene therapy is the treatment of diseases based on the transfer of genetic information. Agents that carry or deliver DNA to target cells are called vectors (Latin vector: carrier, deliverer). Ideally, a vector should accommodate an unlimited amount of inserted DNA, lack the ability of autonomous replication of its own DNA, be easily manufactured, and be available in concentrated form. Secondly, it should have the ability to target specific cell types or to limit its gene expression to specific cell types, and to achieve sustained gene expression in the long term or in a controlled fashion. Finally, it should not be toxic or immunogenic. Such a vector does not exist and none of the DNA delivery systems so far available for in vivo gene transfer is perfect with respect to any of these points. Gene therapy and the means to promote it depend heavily on the development and improvement of new gene vector systems.

A high-copy T7 Escherichia coli expression vector for the production of recombinant proteins with a minimal N-terminal His-tagged fusion peptide

We report here the construction of a vector derived from pET3-His and pRSET plasmids for the expression and purification of recombinant proteins in Escherichia coli based on T7 phage RNA polymerase. The resulting pAE plasmid combined the advantages of both vectors: small size (pRSET), expression of a short 6XHis tag at N-terminus (pET3-His) and a high copy number of plasmid (pRSET). The small size of the vector (2.8 kb) and the high copy number/cell (200-250 copies) facilitate the subcloning and sequencing procedures when compared to the pET system (pET3-His, 4.6 kb and 40-50 copies) and also result in high level expression of recombinant proteins (20 mg purified protein/liter of culture). In addition, the vector pAE enables the expression of a fusion protein with a minimal amino-terminal hexa-histidine affinity tag (a tag of 9 amino acids using XhoI restriction enzyme for the 5'cloning site) as in the case of pET3-His plasmid and in contrast to proteins expressed by pRSET plasmids (a tag of 36 amino acids using BamHI restriction enzyme for the 5'cloning site). Thus, although proteins expressed by pRSET plasmids also have a hexa-histidine tag, the fusion peptide is much longer and may represent a problem for some recombinant proteins.

Modulation of the expression of the transcription factor Max in rat retinal ganglion cells by a recombinant adeno-associated viral vector

Exclusion of the transcription factor Max from the nucleus of retinal ganglion cells is an early, caspase-independent event of programmed cell death following damage to the optic axons. To test whether the loss of nuclear Max leads to a reduction in neuroprotection, we developed a procedure to overexpress Max protein in rat retinal tissue in vivo. A recombinant adeno-associated viral vector (rAAV) containing the max gene was constructed, and its efficiency was confirmed by transduction of HEK-293 cells. Retinal ganglion cells were accessed in vivo through intravitreal injections of the vector in rats. Overexpression of Max in ganglion cells was detected by immunohistochemistry at 2 weeks following rAAV injection. In retinal explants, the preparation of which causes damage to the optic axons, Max immunoreactivity was increased after 30 h in vitro, and correlated with the preservation of a healthy morphology in ganglion cells. The data show that the rAAV vector efficiently expresses Max in mammalian retinal ganglion cells, and support the hypothesis that the Max protein plays a protective role for retinal neurons.

Production of L1 protein from different types of HPV in Pichia pastoris using an integrative vector

Human papillomavirus (HPV) infection is the most common sexually transmitted disease in the world and is related to the etiology of cervical cancer. The most common high-risk HPV types are 16 and 18; however, the second most prevalent type in the Midwestern region of Brazil is HPV-33. New vaccine strategies against HPV have shown that virus-like particles (VLP) of the major capsid protein (L1) induce efficient production of antibodies, which confer protection against the same viral type. The methylotrophic yeast Pichia pastoris is an efficient and inexpensive expression system for the production of high levels of heterologous proteins stably using a wild-type gene in combination with an integrative vector. It was recently demonstrated that P. pastoris can produce the HPV-16 L1 protein by using an episomal vector associated with the optimized L1 gene. However, the use of an episomal vector is not appropriate for protein production on an industrial scale. In the present study, the vectors were integrated into the Pichia genome and the results were positive for L1 gene transcription and protein production, both intracellularly and in the extracellular environment. Despite the great potential for expression by the P. pastoris system, our results suggest a low yield of L1 recombinant protein, which, however, does not make this system unworkable. The achievement of stable clones containing the expression cassettes integrated in the genome may permit optimizations that could enable the establishment of a platform for the production of VLP-based vaccines.