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FERRAZ, ADRIANA C.P.; DALLAVECCHIA, DANIELE L.; SILVA, DÉBORA C.; SILVA- FILHO, RENATO G.; AGUIAR, VALÉRIA M. Post-embriononic development of putoria (Diptera: ) on a diet containing ampicillin in different concentrations Anais da Academia Brasileira de Ciências, vol. 88, núm. 1, marzo, 2016, pp. 105-116 Academia Brasileira de Ciências Rio de Janeiro, Brasil

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Post-embriononic development of Chrysomya putoria (Diptera: Calliphoridae) on a diet containing ampicillin in different concentrations

ADRIANA C.P. FERRAZ1, DANIELE L. DALLAVECCHIA1,2, DÉBORA C. SILVA3, RENATO G. SILVA-FILHO1 and VALÉRIA M. AGUIAR1,2

1Universidade Federal do Estado do Rio de Janeiro, Departamento de Microbiologia e Parasitologia, Laboratório de Estudo de Dípteros, Rua Frei Caneca, 94, Centro, 20211-040 Rio de Janeiro, RJ, Brasil 2Programa de Pós-Graduação em Ciências Biológicas (Biodiversidade Neotropical), Universidade Federal do Estado do Rio de Janeiro, Av. Pasteur, 296, Urca, 22290-240 Rio de Janeiro, RJ, Brasil 3Universidade Estadual do Sudoeste da Bahia, Departamento de Estudos Básicos e Instrumentais, Estrada do Bem Querer, Km 04, 45083-900 Vitória da Conquista, BA, Brasil

Manuscript received on September 1, 2014; accepted for publication on January 16, 2015

ABSTRACT Here we evaluate the effects of different concentrations of the antibiotic ampicillin on the growth and development of Chrysomya putoria. Third-generation, fi rst larvae (L1) reared on 60 grams of homogenate+agar 65% were treated with ampicillin sodium. The experiment consisted of four replicates (40 larvae/replicate) of each antibiotic concentration tested (T1: 466μg/mL ; T2: 81.33 mg/mL and T3: 166.66mg/mL) and a T4: control. The body mass of the mature larvae, after they abandoned the diet, were recorded in batches of fi ve. The variation between the mean body mass of larvae and the duration of larval and pupal stages, and overall duration of the development, viability and normal rates were analyzed by ANOVA. There were no signifi cant differences between the four treatments in the following parameters: body mass of larvae that discontinued the diet as well as the duration of larval, pupal, and total development. The sex ratios found in the four treatments did not differ from those expected. Normality rates were 100% for all treatments. There were no signifi cant differences between treatments for larval and overall viability, but pupal viability differed signifi cantly between T1 and the control, T1 and T2, and between the control and T3. The antibiotic did not appear to signifi cantly alter the development of C. putoria. Key words: Ampicillin, biology, bowfly, Callihoridae, .

INTRODUCTION used pharmaceutical group in self-medication (Mestanza and Pamo 1992). Antibiotics are considered a universal panacea. They account for 12 % of all outpatient prescriptions, Penicillins are the antibiotics of choice to treat however two thirds of all antibiotics are sold a number of pathologies. Ampicillin is indicated without a prescription (Wannmacher 2004). For for the treatment of various infections that are some time, antibiotics have been the most widely caused by gram - positive and gram-negative bacteria: infections of the urinary, respiratory, and Correspondence to: Valéria Magalhães Aguiar E-mail: [email protected] / [email protected] digestive tracts, as well as gall infections, localized

An Acad Bras Cienc (2016) 88 (1) 106 ADRIANA C.P. FERRAZ et al. or systemic infections and infections of the mouth, can alter the rate of development of some species caused by teeth extractions and other surgical of fl ies that feed on cadaver tissues (Gosselin et al. procedures (Porto 2012, Tavares 2014). 2011). Many substances as drugs, or their metabolites, Chysomya putoria has rapid dispersion and can affect the development of of forensic rapid larval development (Oliveira et al. 2007, importance when present in the cadavers that they Ferraz et al. 2011a, 2012). It was originally found colonize (Introna et al. 2001). Substances such in Africa, the Mediterranean and the Middle as scopolamine, buscopan, malathion, diazepam East and was introduced in Brazil in the 1970s. and methadone decrease the development rate In addition, it has high synanthropy and can be of (Carvalho et al. 2001, 2012, Grella et found in several biotopes, including rural, urban al. 2007, Liu et al. 2009, Gosselin et al. 2011). and anthropic forest environments (Ferraz et al. In addition, increased larval development was 2009, Batista-da-Silva et al. 2010, Kosmann et al. observed when insects were exposed to heroin, 2013, Cabrini et al. 2013). This species has been methamphetamine, codeine and paracetamol (Goff reported to cause secondary in and et al. 1991, Carvalho et al. 2004, O`Brien and humans (Guimaraes and Pavavero 1999). It also Turner 2004). However, the effect of penicillin on plays a role as a mechanical vector of pathogenic development remains unknown. organisms (Furlanetto et al. 1984) because of its Insects can even be used as an alternative habit of breeding in decomposing organic matter. method for the detection of toxic substances C. putoria is also known for its potential use in because they do not always metabolize the drugs larval therapy, which is a technique that uses fl y they ingest, leaving traces that can be detected long larvae to remove necrotic tissue and assist in wound after the death of an organism (Hédouin et al. 1999, healing (Dallavecchia et al. 2011). An antibiotic Introna et al. 2001, Carvalho et al. 2001, El-bar and treatment, including penicillin therapy, cannot be Sawaby 2011). interrupted in some patients, however, they can also Insects of forensic importance such as blow undergo larval therapy simultaneously. Worldwide, fl ies are used to estimate the post mortem interval doctors have suggested larval therapy as the fi rst- (IPM) in legal investigations where the time of line treatment for the meticulous removal of dead death is greater than 72 h and the temperature tissue, particularly in diabetic patients. As of the of the cadaver can no longer be used for such 1990s, the Food and Drug Administration (FDA) estimates. The post mortem interval is determined has approved the medicinal use of larvae as an by collecting immature larvae from the body effective treatment for the removal of devitalised which allows the insect to be identified, along tissue in poorly healing wounds (Dallavecchia et with its size and stage (Parry et al. 2011). of al. 2014). the Calliphoridae family should be considered for The species of Calliphoridae can feed on the calculation of the post-mortem interval (PMI) human tissues during their larval stage, either on (Vanlaerhoven 2008). In particular, Chrysomya dead tissues, (participating in the process of corpse species should be considered because they are the decomposition), on live tissues, (causing myiasis) fi rst invaders of carcasses, playing an important or during larval therapy. Thus, these species can role in the sciences (El-bar and Sawaby 2011, absorb drugs from host tissues, which can lead to Oliveira-Costa 2011, Oliveira and Vasconcelos changes in their development rate. Considering 2010). Ignoring toxicological analyses when patients who undergo an antibiotic regimen or calculating PMI can lead to erroneous estimates cases of overuse of antibiotics, and the fact that because certain substances ingested before death we know little about how and for how long

An Acad Bras Cienc (2016) 88 (1) Chrysomya putoria ON A DIET CONTAINING AMPICILLIN 107 larvae accumulate these drugs in their system, mg/mL. The concentrations were chosen from the how they eliminate them and how drugs affect serum ampicillin concentration (after intravenous their development (Soto 2008), we endeavored to dose of 1 g, the maximum concentration of investigate the effects of ampicillin on the post- approximately 40 μg/mL is reached) (Ampicillin embryonic development of Chrysomya putoria 2011). The fi rst concentration chosen (T1) is close (Wiedemann, 1830). to the intravenous serum concentration; the second (T2) is close to the maximum intravenous serum MATERIALS AND METHODS concentration; the third (T3) is approximately Flies were reared and all the experiments were four times the maximum serum intravenous conducted at the Laboratory for the Study of concentration. Each antibiotic concentration tested, Diptera (LED), Department of Microbiology and and the control, were replicated four times using Parasitology, Universidade Federal do Estado do 40 larvae in each repetition. The control received Rio de Janeiro (UNIRIO). distilled water instead of the antibiotic. Adult blowfl ies (C. putoria) were obtained from Dilutions of a commercial injectable ampicillin a stock colony originated from insects collected (ampicillin sodium 1 g; Teuto Lab) were prepared in at the Rio de Janeiro Zoo, which is located at the water for injectable preparations, and 1 mL aliquots Quinta da Boa Vista Park, São Cristóvão, RJ. We were added to the feed to obtain fi nal concentrations used three traps following the model of Mello et al. of 41.6, 81.33 and 166.66 μg/g. The concentration (2007), containing sardine as bait. Traps remained of the antibiotic in the feed was based on the value exposed for about fi ve hours in the morning. After of the serum concentration after the intravenous being collected, adults and larvae of muscoid fl ies injection of 1 g (approximately 40 μg/mL). were taken to the LED where they were sorted and Each beaker was put into a larger beaker (400 identifi ed as per Mello (2003). The fl ies were reared mL) containing sterilized sawdust and was sealed in plastic cages (40x30x20cm) with an opening with elastic escaline. The sawdust was provided in at the top for aeration and an anterior opening to order to allow the larvae to pupate after abandoning allow access to the inside of the cage, which was the diet. The larvae were kept in a climatic chamber covered with escaline coated fabric. The fl ies were at 30 °C/day and 28 °C/night, 70 + 10 % RH and offered water, honey and water (50 %) solution, 14 h photoperiod. Observations were made daily, and chicken gizzard or beef as a protein source, always at the same time (12 h). substrate for oviposition and ovary maturation. The body mass of larvae that abandoned the The rearing methodology followed Barbosa et al. diet was recorded in batches of fi ve with the help (2004) and Ferraz et al. (2011a). of an analytical scale. Larvae were then stored First-instar larvae of the 3rd laboratory in test tubes sealed with nylon and elastic fabric generation were transferred with the help of a fi ne for further observations. We recorded the dates of paintbrush to 100 mL beakers containing 60 g pupation and emergence, as well as the sex ratio gizzard/agar 65 % homogenate (Ferraz et al. 2012). This diet was selected because it is practical and and morphological abnormalities of the adults, sterile (autoclaved), allowing the antibiotic to act when present. fully and only on the larvae. We used 1 g of the The following variables were evaluated: antibiotic generic ampicillin sodium (Teuto Lab). body mass of larvae that discontinued the diet; Each repetition received 1 mL of the antibiotic, duration of post-embryonic development from resulting in the following three fi nal concentrations larval hatching to the interruption of the diet in the food: 41.66 mg/mL, 81.33 mg/mL and 166.66 (period from the inoculation of L1 in the diet

An Acad Bras Cienc (2016) 88 (1) 108 ADRIANA C.P. FERRAZ et al. until the development of third instar larvae, which We used the program Microsoft Excel to ana- tend to abandon the diet in favour of the pupation lyze the raw data and other analyses were conducted substrate); larval stage (the period between the in the program PAST. Variations in the mean body interruption of diet until pupal formation); pupal mass of larvae, the duration of the stages of post- stage (from pupae until the emergence of adults) embryonic development and the viability and and total duration (from L1 to the emergence of normal rates were compared by ANOVA (5 % signi- adults). The frequency of interruption of the larval fi cance). The observed sex ratio was tested against 2 diet, pupation and emergence of adults as well the expected frequency, using chi-square (χ ). as the frequency of male and female emergences RESULTS was graphically demonstrated. We also showed larval, pupal and total viability, gender ratio and The four treatments did not differ signifi cantly with percentage of normal insects (i.e. those with no respect to the body mass of C. putoria larvae (Table visible morphological deformities such as lack of I). Furthermore, the average duration from L1 distention of the wings, visible reduction of body until the discontinuation of the diet, larval, pupal size, wrinkled cuticles, sharp increase in the head and total (days) stages did not differ signifi cantly size and retracted pitilinum). among the four treatments (Table II).

TABLE I Body mass of larvae (g)* of the blowfl y Chrysomya putoria (Wiedemann, 1830) (Diptera, Calliphoridae) after four treatments with different concentrations of ampicillin, reared in a climatic chamber (30 °C day and 28 °C night, 70 + 10 % RH and 14 h photoperiod). Mean individual mass (g) ± Treatment Variation interval Standard deviation Control 0.039a ± 0.015 0.014 - 0.059 T1 0.038a ± 0.012 0.012 - 0.058 T2 0.049a ± 0.008 0.016 - 0.062 T3 0.046a ± 0.005 0.031 - 0.056 * Means followed by same letter in same column do not differ signifi cantly by ANOVA, 5 % confi dence. control = gizzard homogenate agar; T1 = gizzard homogenate agar + ampicillin 41.66 μg/mL, T2 = gizzard homogenate agar + ampicillin 81.33 μg/mL, T3 = gizzard homogenate agar + ampicillin 166.66 μg/mL.

TABLE II Average duration of the post-embryonic development stages *(days) of larvae of the blowfl y Chrysomya putoria (Wiedemann, 1830) subjected to four treatments with different concentrations of ampicillin and reared in a climatic chamber (30 °C day and 28 °C night, 70 + 10 % RH and 14 h photoperiod). Variation Standard Day interval Deviation Control 3.617a 3 - 4 0.466 T1 3.510a 3 - 4 0.412 T2 3.159a 3 - 4 0.177 abandonment Duration until T3 4.498a 3 - 6 2.477

An Acad Bras Cienc (2016) 88 (1) Chrysomya putoria ON A DIET CONTAINING AMPICILLIN 109

TABLE II (continuation) Variation Standard Day interval Deviation Control 4.617a 4 - 5 0.466 T1 4.016a 4 - 5 0.729 T2 4.174a 4 - 5 0.174

Larval stage T3 4.469a 4 - 7 0.700 Control 3.660a 3 - 5 0.271 T1 3.664a 3 - 5 0.490 T2 3.807a 3 - 5 0.170

Pupal stage T3 3.623a 3 - 5 0.420 Control 8.279a 8 - 9 0.244 T1 8.183a 8 - 9 0.223

Total Total T2 7.888a 8 - 10 0.333 duration T3 8.218a 8 - 9 0.195 *Means followed by same letter in same column do not differ signifi cantly by ANOVA, 5 % confi dence. Control = gizzard homogenate in agar; T1 = gizzard homogenate in agar + 41.66 μg/ mL ampicillin; T2 = gizzard homogenate in agar + 81.33 μg/mL ampicilin, T3 = gizzard homogenate in agar + 166.66 μg/mL ampicillin.

The rates of diet abandonment, pupation and (Table III). The pupal viability of T1 (lower than emergence (%) obtained for C. putoria in the four in the other treatments) differed signifi cantly from different treatments are shown in Figure 1. There is the control and T2, and the pupal viability of T3 very little difference in the peaks of abandonment, differed from the control. pupation and emergence between the four treatments. DISCUSSION The sex ratios were as follows: control (males Human myiasis is the presence of fl y larvae in human = 48 %, females = 52 %); T1 (males = 48 %, tissues, where they feed and develop as parasites. females = 52 %), T2 (males = 54 %, females = 46 They preferentially develop in the human skin but %) and T3 (males = 49 % females = 51 %). The frequently reach natural body cavities (Ferraz et al. chi-square tests revealed that the sex ratios found in 2011b). They constitute an important public health the four treatments did not differ from the expected problem because of the strong social component 2 2 2 (control: χ = 0.001, T1: χ = 0.001, T2: χ =0.007, associated with their presence. Myiasis can occur 2 2 T3: χ = 0.001; tabulated χ = 3.84, gl = 1, α = 5 %). in areas previously affected by other illnesses All adults (100 %) in the control and in the (or dermatoses), including erysipelas, which are other treatments were normal, i.e., showed no treated with penicillin as the fi rst option in most morphological deformities. The rate of emergence cases (Braga et al. 2011). This pathology is caused of males and females of C. putoria was similar by species of the Calliphoridae family, mainly (Fig. 2), the emergence peak of both occurred on C. putoria, which has scavenger/necrophagic the 8th day. Only in T2 and T3 the emergence of behaviour and develops in dead tissues (corpses) females extended up to the 10th day. or necrotic wounds of living hosts (Guimaraes and As for larval and overall viability, there were Pavavero 1999). It is known that certain chemicals no signifi cant differences between the treatments and hormones exert a deleterious effect on insect

An Acad Bras Cienc (2016) 88 (1) 110 ADRIANA C.P. FERRAZ et al.

Abandonment Pupated Emerged

Figure 1 – Rate of diet abandonment, pupation and emergence (%) of Chrysomya putoria (Wiedemann, 1830) subjected to four treatments: control = gizzard homogenate agar; T1 = gizzard homogenate agar + ampicillin 41.66 μg/mL, T2 = gizzard homogenate agar + ampicillin 81.33 μg/mL, T3 = gizzard homogenate agar + ampicillin 166.66 μg/mL. development (Carvalho et al. 2001, 2012, Liu et al. 2011). At present, larval therapy is used in medical 2009, Gosselin et al. 2011). However, the effect of procedures in over 30 countries worldwide. On the antibiotics such as ampicillin on the development other hand, strategies for the sterilisation of eggs of fl y larvae remains unknown. and disinfection of larvae are still being developed Blowfly larvae (Calliphoridae) can be used in Brazil (Dallavecchia et al. 2014). Antibiotic in larval therapy for treating different types of therapy is often associated with larval therapy; wounds. These larvae remove the necrotic tissue, however, the extent to which antibiotics can affect stimulate granulation tissue and promote microbial the development of larvae used for medicinal decontamination (Dallavecchia et al. 2011). purposes remains unknown. Lucilia sericata larvae (Meigen, 1826) (Diptera: Another fi eld of biology in which calliphorid Calliphoridae) are the most commonly used fl ies are important is forensic entomology, which species in the Northern Hemisphere, whereas in studies the biology and behaviour of insects and Brazil, Chrysomya species have the potential of other arthropods during criminal investigations. being used for larval therapy (Dallavecchia et al. The most common use of this knowledge is for

An Acad Bras Cienc (2016) 88 (1) Chrysomya putoria ON A DIET CONTAINING AMPICILLIN 111

Figure 2 – Rate of emergence, in days, of males and females of Chrysomya putoria (Wiedemann, 1830) subjected to four treatments: control = gizzard homogenate agar; T1 = gizzard homogenate agar + ampicillin 41.66μg/mL, T2 = gizzard homogenate agar + ampicillin 81.33 μg/mL, T3 = gizzard homogenate agar + ampicillin 166.66 μg/mL.

TABLE III ANOVA comparison of the viability of larvae, pupae and total (L1 to adult) of Chrysomya putoria (Wiedemann, 1830) subjected to four treatments: control = gizzard homogenate agar; T1 = gizzard homogenate agar + ampicillin 41.66 μg/mL, T2 = gizzard homogenate agar + ampicillin 81.33 μg/ mL, T3 = gizzard homogenate agar + ampicillin 166.66 μg/mL. p= signifi cance value (ANOVA) Viability*(%) Control T1 T2 T3 Control 82.50a - 0.176 0.444 0.492 T1 96.88a 0.176 - 0.162 0.113 T2 90.63a 0.444 0.162 - 0.214 Larval T3 70.00a 0.492 0.113 0.214 - Control 96.37a - 0.024 0.929 0.008 T1 78.46b 0.024 - 0.048 0.371

Pupal T2 95.95a 0.929 0.048 - 0.056 T3 84.42b 0.008 0.371 0.056 -

An Acad Bras Cienc (2016) 88 (1) 112 ADRIANA C.P. FERRAZ et al.

TABLE III (continuation) p= signifi cance value (ANOVA) Viability*(%) Control T1 T2 T3 Control 80.00a - 0.764 0.560 0.223 T1 76.25a 0.764 - 0.237 0.246

Total T2 86.88a 0.560 0.237 - 0.072 T3 58.75a 0.223 0.246 0.072 - *Means followed by same letter in same column do not differ signifi cantly by ANOVA, 5 % confi dence. determining PMI, which estimates the period for tested corresponded to an overdose of antibiotics, which the body remained at the crime scene and equivalent to concentrations that were two to four the period of death. By providing an estimates times greater than the serum concentration. In the of PMI, particularly between 2 and 4 weeks of present study, a few parameters were altered by the decomposition, it is assumed that insects will presence of ampicillin, as will be shown below. develop at predictable rates, considering certain To evaluate the effects of different substances environmental conditions. Moreover, the effects of on fly development, many authors have used drugs on insect development need to be considered guinea pig tissues (Carvalho et al. 2001, George (Oliveira-Costa 2011). Ignoring this data may et al. 2009, Liu et al. 2009) in addition to natural result in errors in the PMI estimates of up to days diet along with the substance to be tested (Grella (Parry et al. 2011). et al. 2007, Parry et al. 2011). However, artifi cial Antibiotics are commonly used to cure diets can be used to homogeneously introduce an established infection and have the effect of substances in treatments, dispensing with the need hindering or preventing bacterial growth (Porto for cobaios, which are expensive to handle (Soto 2012). Therefore, patients with myiasis, who often 2008). Similarly, we know the exact composition have infected wounds, or patients who make use of artifi cial diets, and by using them we are able of larval therapy, usually take this type of drug. to prevent possible unwanted interactions with the Considering also that the number of illicit drug substance to be tested. The artifi cial diet used in users, and users of other toxic substances, have our study allowed the consumption of ampicillin gradually increased over the years, studies on at different concentrations, to observe potential entomotoxicity are important to understand the reactions of fl ies in the presence of this compound possible interactions between insects and these in their diet. Moreover, this diet contains few substances (Introna et al. 2001, UNODC 2007). ingredients, which is an advantage when the When scavenger insects ingest drugs by feeding influence of chemicals on insect development on the corpses of individuals who used drugs, their needs to be evaluated. The agar base used in this rate of development and metabolism may be altered artifi cial diet has been used in the formulation of (Introna et al. 2001). We tested the effect of three diets for other insect groups, and most of them concentrations of ampicillin on the development are with insects of agricultural importance (Parra of C. putoria. The concentrations chosen were 2002). similar to serum intravenous concentrations in Previous studies have shown that, in callipho- cases in which individuals were taking ampicillin rids, adults end up smaller and lighter when there according to medical recommendations for the are limitations in space and food during their treatment of infections; the other concentrations development. Consequently, the fi nal size of adults

An Acad Bras Cienc (2016) 88 (1) Chrysomya putoria ON A DIET CONTAINING AMPICILLIN 113 refl ects the quantity and quality of the nutrition the results between the first and second studies consumed during the larval stage (Goodbrod and and ours did not exceed one day. There were Goff 1990, Reis et al. 1994). Considering that the also no signifi cant differences in the duration of body mass of larvae was not signifi cantly different larval, pupal, and total development between the among the four treatments, we can conclude that control and each of the treatments with ampicillin. a decrease in food intake and/or food absorption, Therefore, it is possible that the antibiotic did even in the presence of antibiotics, did not take not affect the conversion of food, i.e. during all place. The diet we provided was rich in proteins and treatments, larvae reached similar body weights is adequate for calliphorids (Yaghoobi et al. 2005), when feeding on the substrate in the same period. which seek to ingest as much food as possible Larvae in T3, which received the highest before the resources are exhausted. The larvae then antibiotic concentration, took a day longer to disperse in the search for new food sources or a abandon the diet, pupate, and emerge than the location to pupate (Greenberg 1990), which also other treatments. It is possible that the antibiotic happened in the present study when they switched changed some organoleptic characteristics of their to the larger beaker containing sawdust. diet. It has been shown that the behaviour of C. The amount of gizzard homogenate + agar albiceps (Diptera: Caliphoridae) larvae is altered used in the diet in each repetition also secured by palatability and by pheromones (Faria et al. the ideal relative density of more than 1 g of 2004). The presence of drugs in tissues can affect meat per , as advocated by Aguiar-Coelho larval development of scavenger fl ies that feed on and Milward-de-Azevedo (1996) for different these substrates. In many cases, PMI is calculated calliphorids, as a means to avoid the stress of by measuring the weight and age of immature exploitative competition for resources or chemical larvae collected in a body; therefore, ignoring this changes in the food substrate resulting from the interfering factor may lead to an underestimation of larval metabolism (Khazaeli et al. 1993, Bubli et the time of death (up to 48 h) (Thyssen and Grella al. 1998). The amount of food can also alter the 2011). phenotype (Ribeiro et al. 1993, Reis et al. 1994, Nevertheless, the population of flies that Roper et al. 1996, Ribeiro 1998), but even in the emerged from all treatments can be considered treatments with ampicillin, 100 % of the adults stable, since none diverged from the expected were normal. sex ratio 1:1 and the rate of emergence of males Oliveira et al. (2007), in a study using C. and females was also similar (only in T2 and T3 putoria reared on beef (acclimatized chamber at 27 the emergence of females extended up to one ºC ± 1, 60 + 10 % RH. and 14 photophase), observed day longer). Viability was also above 60 % in all that larval development lasts 92 h (approximately treatments, except for total viability in T3. This 3.83 days). This is less than the durations obtained value is considered promising in Diptera (Loureiro in our four treatments. Ferraz et al. (2011a), using et al. 2005) for laboratory studies with diets. beef and dog food at room temperature (average Despite the decreased viability of fl ies that fed on max T = 25.0 ºC, mean average T = 21.5 ºC, RH 60- the diet with the highest antibiotic concentration, 90 %) found higher values. The increase in the time on average, 60 % of the immature stages developed it takes for the larval stage to complete, is a strategy into the adult stage; therefore, this decrease was not to offset the decline in the nutritional quality of signifi cant. the food consumed (Aguiar-Coelho and Milward- Some authors believe that there is a correlation de-Azevedo 1996). However, the differences in between the concentration of drugs present in the

An Acad Bras Cienc (2016) 88 (1) 114 ADRIANA C.P. FERRAZ et al. tissues eaten by larvae, and the concentration of RESUMO the same drugs in the larval tissues (Introna Jr et al. Nós avaliamos aqui os efeitos de diferentes concen- 1990, Kintz et al. 1990). Other authors, by contrast, trações do antibiótico ampicilina sobre o crescimento indicate that no such relationship exists, because e desenvolvimento de Chrysomya putoria. the process of accumulation and elimination of de primeiro instar (L1), de terceira geração, criadas substances by the larvae may vary according to the em 60 gramas de homogenato de moela em agar 65% drug and the species analyzed (Sadler et al. 1997). foram tratadas com ampicilina sódica. O experimento In skeletonised bodies or those at an advanced stage consistiu de quatro repetições (40 larvas / réplicas) para of decomposition, where traditional sources of cada concentração do antibiótico testado (T1: 66μg/ mL; T2: 81,33μg/mL e T3: 166,66μg/mL) e o T4: analysis such as blood, urine or internal organs are controle. A massa corporal das larvas maduras, após not available, insects may serve as reliable sources abandonarem a dieta, foi registrada em lotes de cinco. for toxicological analyses. In addition, insects are A variação entre a massa corporal média de larvas e a used in the detection of licit or illicit drugs present duração das taxas de desenvolvimento de larva, e in the body, including stimulants, CNS depressants, a duração total do desenvolvimento, viabilidades e taxas anti-depressants, analgesics, amphetamines and de normalidade foram analisadas por ANOVA. Não cocaine. Entomotoxicological studies should houve diferença signifi cativa entre os quatro tratamentos primarily focus on drug bioaccumulation and nos seguintes parâmetros: massa corporal de larvas que abandonaram a dieta, bem como, a duração larval, metabolism of necrophagic insects as well as drugs pupal e desenvolvimento total. As proporções sexuais and poisons concentrations or their metabolites in encontradas nos quatro tratamentos não diferiram do insect and human tissues (Introna Jr et al. 1990). esperado. Taxas de normalidade foram de 100% para If we knew the different ways by which todos tratamentos. Não houve diferença signifi cativa absorption, bioaccumulation and excretion of subs- entre os tratamentos para as viabilidade larval e total, tances/drugs take place in fl y larvae, this could help mas a viabilidade de pupa diferiu signifi cativamente us understand some of our contradictory results, for entre T1 e o controle, T1 e T2, e entre o controle e T3. instance the signifi cant difference in the viability of O antibiótico pareceu não alterar signifi cativamente o desenvolvimento de C. putoria. T1 with respect to the control and T2, and the pupal Palavras-chave: Ampicilina, biologia, moscas varejeiras, viability of T3 differing from the control. Calliphoridae, entomologia forense. Few differences were observed after the introduction of the antibiotic ampicillin in the REFERENCES larval diet, therefore its presence in the food source AGUIAR-COELHO VM AND MILWARD-DE-AZEVEDO in the concentrations used in this study does not EMV. 1996. Relações intra-específicas de Cochliomyia signifi cantly interfere with larval development. macellaria (Fabricius) e (Wiedemann), (Fabricius) ACKNOWLEDGMENTS e Cochliomyia macellaria (Fabricius) (Calliphoridae, Diptera) sob condições de laboratório. Rev Bras Entomol We thank Conselho Nacional de Desenvolvimento 41: 35-40. Científico e Tecnológico (CNPq), Financiadora BARBOSA LS, JESUS DML AND AGUIAR-COELHO VM. 2004. Longevidade e capacidade reprodutiva de casais agrupados de Estudos e Projetos (FINEP), Universidade de Chrysomya megacephala (Fabricius, 1794) (Diptera: Federal do Estado do Rio de Janeiro (UNIRIO) Calliphoridae) oriundos de larvas criadas em dieta natural and Fundação Carlos Chagas Filho de Amparo à e oligídica. Rev Bras Zoociências 6: 207-217. BATISTA-DA-SILVA JA, MOYA-BORJA GE AND QUEIROZ Pesquisa do Estado do Rio de Janeiro (FAPERJ) for MMC. 2010. Ocorrência e Sazonalidade de Muscóides the fi nancial support for this work. (Diptera,Calliphoridae) de Importância Sanitária no

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