Detection of terbufos in cases of intoxication by means of entomotoxicological analysis using ATR-FTIR spectroscopy combined with chemometrics.

The detection of toxic substances in larvae from carcasses in an advanced stage of decomposition may help criminal expertise in elucidating the cause of death in suspected cases of poisoning. Terbufos (Counter®) or O,O-diethyl-S-[(tert-butylsulfanyl)methyl] phosphorodithioate is an insecticide and systemic nematicide, which has very high toxicity from an acute point of view (oral LD50 in rodents ranging from 1.4 to 9.2 mg/kg) that has been marketed irregularly and indiscriminately in Brazil as a rodenticide, often being used to practice homicides. The present study aims to evaluate the use of attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy to detect traces of terbufos pesticide in fly larvae (Sarcophagidae). ATR-FTIR spectra of scavenger fly larvae from control (n = 31) and intoxicated (n = 80) groups were collected and submitted to chemometric analysis by means of multivariate classification using principal component analysis with quadratic discriminant analysis (PCA-QDA), successive projections algorithm with quadratic discriminant analysis (SPA-QDA) and genetic algorithm with quadratic discriminant analysis (GA-QDA) in order to distinguish between control and intoxicated groups. All discriminant models showed sensitivity and specificity above 90%, with the GA-QDA model showing the best performance with 98.9% sensitivity and specificity. The proposed methodology proved to be sensitive and promising for the detection of terbufos in scavenger fly larvae from intoxicated rat carcasses. In addition, the non-destructive nature of the ATR-FTIR technique may be useful in preserving the forensic evidence, meeting the precepts of the chain of custody and allowing for counter-proof.

Near infrared spectroscopy (NIRS) coupled with chemometric methods to identify and estimate taxonomic relationships of flies with forensic potential (Diptera: Calliphoridae and Sarcophagidae).

Infrared spectroscopy has been gaining prominence in entomology, such as for solving taxonomic problems, sexing adult specimens, determining the age of immature specimens, detecting drugs of abuse in fly larvae, and can be an important technique in Forensic Entomology. In order to help identify the species of Calliphoridae and Sarcophagidae families, the present study aimed to evaluate the use of near infrared spectroscopy (NIRS) coupled with chemometric methods for separating fly specimens into taxonomic categories and understanding the taxonomic relationship between them. Spectra collected from nine species of flies were subjected to unsupervised principal component analysis (PCA) and hierarchical cluster analysis (HCA), in which we sought to visualize the relationship between the samples (segregation of genera and families) with subsequent identification. In PCA, the best model was achieved using five principal components (PCs), which explained 99.16% of total variance of the original data set. The first principal component (PC1) and the fourth principal component (PC4) provided the best segregation, the latter being more important in the segregation of the species Chrysomya albiceps, Lucilia eximia, and Ravinia belforti from the others. In the HCA dendrogram, there was a clear separation between the specimens by family (Calliphoridae and Sarcophagidae) and genera (Chrysomya, Lucilia, Oxysarcodexia, Peckia and Ravinia). This study shows that NIRS is efficient to identify flies' taxonomic properties, such as family and genera, providing quick evidence for the tested species identity.

Infrared spectroscopy (NIRS and ATR-FTIR) together with multivariate classification for non-destructive differentiation between female mosquitoes of Aedes aegypti recently infected with dengue vs. uninfected females.

One of the most important steps in preventing arboviruses is entomological surveillance. The main entomological surveillance action is to detect vector foci in the shortest possible stages. In this work, near and medium infrared spectra collected from female Aedes aegypti mosquitoes recently infected and not infected with dengue were used in order to build chemometric models capable of differentiating the spectra of each class. For this, computational algorithms such as Successive Projection Algorithm (SPA) and Genetic Algorithm (GA) were used together with Linear Discriminant Analysis (LDA). The constructed models were evaluated with sensitivity and specificity calculations. It was observed that models based on near infrared (NIR) spectra have better classification results when compared to mid infrared (MIR) spectra, as well as models based on GA present better results when compared to those based on SPA. Thus, NIR-GA-LDA obtained the best results, reaching 100.00 % for sensitivity and specificity. NIR spectroscopy is 18 times faster and 116 times cheaper than RT-qPCR. The findings reported in this study may have important applications in the field of entomological surveillance, prevention and control of dengue vectors. In the future, mosquito traps equipped with portable NIR instruments capable of detecting infected mosquitoes may be used, in order to enable an action plan to prevent future outbreaks of the disease.

Myiasis in humans: Case reports in Northeastern Brazil including multispecies co-infestation by Sarcophagidae.

Myiasis is a parasitosis characterized by an infestation of living vertebrates (humans and other animals) by Diptera larvae, whose occurrence and etiological identification are still neglected by health professionals. Here we analyzed the human myiasis cases registered from 2010 to 2018 in health care units in the municipality of Natal, Northeast region of Brazil. Specifically, we aimed to: I) analyze the medical records of cases documented from 2010 to 2017; II) list the patients predisposing factors; and III) monitor the recent cases diagnosed in health units between August 2017 and March 2018 and report the taxonomic identity of the infesting species. Our data revealed that myiases mainly affect the elderly and individuals with predisposing medical conditions (e.g. senility, filariasis). Regarding the new cases, larvae of Calliphoridae (C. hominivorax (Coquerel, 1858)) and Sarcophagidae [Sarcophaga (Liopygia) ruficornis (Fabricius, 1794), Peckia (Sarcodexia) lambens (Wiedemann, 1830), and Helicobia morionella (Aldrich, 1930)] were identified. Furthermore, this was the first report of a co-infestation with three Sarcophagidae species and also the presence of H. morionella in a human wound. These results reinforce that myiasis is an underreported parasitosis, which may underestimate the ability of previously unreported fly species to feed on living human tissues in the Neotropical Region.

Infrared spectroscopy and forensic entomology: Can this union work? A literature review.

For more than two decades, infrared spectroscopy techniques combined with multivariate analysis have been efficiently applied in several entomological fields, such as Taxonomy and Toxicology. However, little is known about its use and applicability in Forensic entomology (FE) field, with vibrational techniques such as Near-infrared spectroscopy (NIRS) and Medium-infrared spectroscopy (MIRS) underutilized in forensic sciences. Thus, this work describes the potential of NIRS, MIRS, and other spectroscopic methodologies, for entomological analysis in FE, as well as discusses its future uses for criminal or civil investigations. After a thorough research on scientific journals database, a total of 33 publications were found in scientific journals, with direct or indirect application to FE, including experimental applications of NIRS and MIRS in taxonomic discrimination of species, larval age prediction, detection of toxic substances in insects from environments or crime scenes, and detection of internal or external infestations by live or dead insects in stored products. Besides, NIRS and MIRS combined with multivariate analysis were efficient, inexpensive, fast, and non-destructive analytical tools. However, more than 51% of the spectroscopic publications are concentrated in the stored products field, and so we discuss the need for expansion and more direct application in other FE areas. We hope the number of articles continues to increase, and as NIRS and MIRS technology progress, they advance in forensic research and routine use.

Differential Ability of Necrophagous Diptera to Colonize Concealed Resources: Empirical Evidence From a Field Experiment in Brazil.

Field studies in forensic entomology seem to assume that a cadaver is a resource equally accessible to flies, irrespective of the species' size, behavior, and sensorial capacity. We aimed to challenge this assumption by experimentally manipulating baited traps in order to test whether the accessibility to decomposing baits is related to the following: (i) the diversity of necrophagous dipterans; (ii) the body size of adults; and (iii) the female sexual maturity stage. The experiment was performed in a rainforest fragment in Northeastern Brazil, in an area characterized by high levels of homicides. Traps baited with decomposing chicken liver had three levels of access: easy (six openings on the trap), intermediate (three), and difficult (one). A total of 750 adults of Calliphoridae, Fanniidae, Muscidae, Phoridae, and Sarcophagidae were collected. Of the 18 species recorded, Megaselia scalaris was the most abundant (46.8%) followed by Lucilia eximia (29.7%) and Chrysomya megacephala (9.8%). The number of species increased significantly with the accessibility level, but overall abundance did not differ, as a consequence of the numerical dominance of M. scalaris. A strong female bias (>91%) was observed in all traps. For Calliphoridae specimens, 90% of the females caught in traps under intermediate and difficult access were sexually mature. By empirically demonstrating that limitation of access and exclusion of visual stimuli do not deter several dipteran species from reaching a substrate, we expand the knowledge on the factors that can shape the colonization of concealed human cadavers, including variables such as insect size and stage of sexual maturity.

A novel use of infra-red spectroscopy (NIRS and ATR-FTIR) coupled with variable selection algorithms for the identification of insect species (Diptera: Sarcophagidae) of medico-legal relevance.

Unequivocal identification of fly specimens is an essential requirement in forensic entomology. Herein, a simple, non-destructive and rapid method based on two vibrational spectroscopy techniques [Near-Infrared Spectroscopy (NIRS) and attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy] coupled with variable selection techniques such as genetic algorithm-linear discriminant analysis (GA-LDA) and successive projection algorithm-linear discriminant analysis (SPA-LDA) were applied for identifying and discriminating six species of flesh flies (Diptera: Sarcophagidae) native to Neotropical regions. This novel approach is based on the unique spectral "fingerprints" of their biochemical composition. One hundred sixty (160) NIRS and FT-IR specimens (120 male, 40 female) were acquired; different pre-processing methods such as baseline correction, derivative and Savitzky-Golay smoothing were also performed. In addition, the multivariate classification accuracy results were tested based on sensitivity, specificity, positive (or precision) and negative predictive values, Youden index, positive and negative likelihood ratios. Principal components analysis (PCA) was employed for male vs. female category using NIRS, strongly showing the separation between the classes with only three principal components and 99% explained variance. Differentiation between the genera Oxysarcodexia, Peckia and Ravinia was efficiently confirmed by both techniques. In comparison with other biological methods, this approach represents an effective choice for fast and non-destructive identification in forensic entomology.

Use of maggot therapy for treating a diabetic foot ulcer colonized by multidrug resistant bacteria in Brazil.

This study reports the efficacy of maggot therapy in the treatment of diabetic foot ulcer infected with multidrug resistant microorganisms. A 74 year old female patient with diabetes for over 30 years, was treated with maggot therapy using larvae of Chrysomya megacephala. The microbiological samples were collected to evaluate aetiology of the infection. The therapy done for 43 days resulted in a reduction of necrosis and the ulcer's retraction of 0.7 cm [2] in area. Analysis of the bacteriological swabs revealed the presence of Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa. Further studies need to be done to confirm the role of maggot therapy in wound healing using a large sample and a proper study design.

[Periodicity of capture of the Anopheles darlingi Root (Diptera: Culicidae) in Porto Velho, Rondônia, Brazil]. / Periodicidade de captura de Anopheles darlingi Root (Diptera: Culicidae) em Porto Velho, RO.

Anopheles darlingi Root is the principal malaria vector in Amazonia region. The objectives of this work were to study the periodicity of Anopheles darlingi Root, the host preference for peri or extra-domestic environments and the parous rate in four field sites in Porto Velho (RO) by human-landing. All of the Anopheles specimens collected were identified, but only A. darlingi was dissected for the parous study. The results showed that human-landing colleted a total of 985 anophelines, with A. darlingi (972) being the most abundant species. Female mosquitoes were more abundant at extra-domestic environments in two of the locations studied (São João e Candeias do Jamari) (P < 0.05). The parous rate was 96% and no periodicity was observed for captures of females of A. darlingi in the field sites.

Periodicidade de captura de Anopheles darlingi Root (Diptera: Culicidae) em Porto Velho, RO / Periodicity of capture of the Anopheles darlingi Root (Diptera: Culicidae) in Porto Velho, Rondônia, Brazil

Anopheles darlingi Root é o principal vetor de malária na Amazônia brasileira. Os objetivos deste trabalho foram avaliar a periodicidade de captura, a preferência por peri ou extradomicílio e a taxa de paridade de A. darlingi em quatro localidades em Porto Velho, RO, utilizando a atração humana. Todos os anofelinos capturados foram identificados e A. darlingi foi dissecada para caracterização da paridade. Do total de 985 anofelinos coletados, 972 eram A. darlingi. O número de fêmeas foi significativamente maior no extradomicílio em duas das localidades estudadas (São João e Candeias do Jamari) (P < 0,05). A taxa de paridade foi de 96 por cento e não foi observado horário preferencial para captura de fêmeas de A. darlingi nas localidades estudadas.

Anopheles darlingi Root is the principal malaria vector in Amazonia region. The objectives of this work were to study the periodicity of Anopheles darlingi Root, the host preference for peri or extra-domestic environments and the parous rate in four field sites in Porto Velho (RO) by human-landing. All of the Anopheles specimens collected were identified, but only A. darlingi was dissected for the parous study. The results showed that human-landing colleted a total of 985 anophelines, with A. darlingi (972) being the most abundant species. Female mosquitoes were more abundant at extra-domestic environments in two of the locations studied (São João e Candeias do Jamari) (P < 0.05). The parous rate was 96 percent and no periodicity was observed for captures of females of A. darlingi in the field sites.

Evaluation of the sticky MosquiTRAP for detecting Aedes (Stegomyia) aegypti (L.) (Diptera: Culicidae) during the dry season in Belo Horizonte, Minas Gerais, Brazil.

MosquiTRAP is a sticky trap specifically designed to capture gravid females of Aedes aegypti (L.) and allows the identification of the mosquito in the field during the inspection of the trap. This study aims to compare this sticky trap to larval and ovitrap surveys for field monitoring of A. aegypti during the dry season. The study was conducted from March to June of 2003 in 20 blocks of the district of Itapoã, Belo Horizonte, MG. The traps were monitored every week while the larval survey was conducted on a monthly basis. The larval index: Premise Index (PI) and Breteau Index (BI) had equal values throughout the experiment (1.72 in the first two months and zero in the last two). The container index (CI) during the first two months was 0.09 and 0.1%, respectively and zero in the last two. The Ovitrap Positive Index (OPI) ranged from 16.7 to 76.9%, and the MosquiTRAP Positive Index (MPI) ranged from 0 to 31.5%. The Egg Density Index (EDI) ranged from 26.6 to 82.8, while the Adult Density Index ranged from 0 to 1.6 throughout the experiment. Temperature and rainfall did not affect the Positive and Density Indices, although these environmental variables seemed to have affected the larvae indices. Although the MosquiTRAP caught a low number of Aedes mosquitoes during the study, it was more sensitive than the larval survey to detect the presence of Aedes mosquitoes.

Evaluation of the sticky MosquiTRAP™ for detecting Aedes (Stegomyia) aegypti (L.) (Diptera: Culicidae) during the dry season in Belo Horizonte, Minas Gerais, Brazil

MosquiTRAP™ is a sticky trap specifically designed to capture gravid females of Aedes aegypti (L.) and allows the identification of the mosquito in the field during the inspection of the trap. This study aims to compare this sticky trap to larval and ovitrap surveys for field monitoring of A. aegypti during the dry season. The study was conducted from March to June of 2003 in 20 blocks of the district of Itapoã, Belo Horizonte, MG. The traps were monitored every week while the larval survey was conducted on a monthly basis. The larval index: Premise Index (PI) and Breteau Index (BI) had equal values throughout the experiment (1.72 in the first two months and zero in the last two). The container index (CI) during the first two months was 0.09 and 0.1 percent, respectively and zero in the last two. The Ovitrap Positive Index (OPI) ranged from 16.7 to 76.9 percent, and the MosquiTRAP Positive Index (MPI) ranged from 0 to 31.5 percent. The Egg Density Index (EDI) ranged from 26.6 to 82.8, while the Adult Density Index ranged from 0 to 1.6 throughout the experiment. Temperature and rainfall did not affect the Positive and Density Indices, although these environmental variables seemed to have affected the larvae indices. Although the MosquiTRAP caught a low number of Aedes mosquitoes during the study, it was more sensitive than the larval survey to detect the presence of Aedes mosquitoes.

MosquiTRAP é uma armadilha desenvolvida para a captura de fêmeas grávidas de Aedes aegypti (L.) e permite a identificação do mosquito em campo, durante a vistoria da armadilha. O objetivo desse estudo foi comparar essa armadilha com a pesquisa larvária e a ovitrampa para o monitoramento do A. aegypti em campo durante a estação seca. O trabalho foi realizado de março a junho 2003 em 20 quarteirões no bairro Itapoã, Belo Horizonte (MG). O monitoramento das armadilhas foi semanal e a pesquisa larvária foi realizada mensalmente. O índice Predial (IP) e o índice de Breteau (IB) apresentaram valores iguais durante todo o experimento (1,72 nos primeiros dois meses e zero nos dois últimos) e o índice de Recipiente (IR) nos dois primeiros meses foi de 0,09 e 0,1 por cento respectivamente e nos dois últimos foi zero. O índice de Positividade de Ovitrampa (IPO) variou de 16,7 a 76,9 por cento e o índice de Positividade de MosquiTRAP (IPM) variou de 0 a 31,5 por cento. O índice de Densidade de Ovos (IDO) variou de 26,6 a 82,8 enquanto o índice de Densidade de Adultos (IDA) variou de 0 a 1,6 durante todo o experimento. A temperatura e a precipitação não influenciaram os índices de Positividade e de Densidade, mas parecem ter influenciado os índices larvários. Apesar de a MosquiTRAP ter coletado um número pequeno de Aedes, observou-se que ela foi mais sensível para detectar a presença de Aedes do que a pesquisa larvária.