Chronometrical Morphology Changes Of Larval Of Three Species Necrophagous Flies And Their Valuation In Forensic Medicine

Objective: To study the growth and development patterns of the cephalopharyngeal bone of Aldrichina grahami,Chrysomya megacephala and Lucilia sericata larval at different temperatures and accumulate the developmental biology data on necrophagous flies dominated in Shijiazhuang area, which would provide scientific database for deducing the postmortem interval (PMI) in criminal investigation.Methods:Adult flies of Chrysomya megacephala,Lucilia sericata and Aldrichina grahami were collected outdoors. The experiment was performed on the third generation of offsprings. They were reared successively in different biochemical incubators at the temperature of 20℃, 24℃, 28℃respectively. The humidity, photoperiod and food supply were kept unchanged during the incubation. 10 larval were randomly collected from each group from larvae being laid until the beginning of the prepuparial stage at 12 intervals . The larvae were fixed in boiling water and then preserved in 70% alcohol mixed with glycerine. The samples were taken out from the preserving solution and the liquid of surface was blotted with filter paper. Some holes were made on the abdomens of specimen with a pin, then put them in 10% KOH for 24 hours to corrod their innards. The larval were pressed gently to let the corroded tissues excreted. And then washed several times with clean water, two fragment of larval body were cut out for morphological observation under stereomicroscope, the fragments between scolex section and the second thoracic section were used for preparing specimens of anterior spiracle and cephalopharyngeal skeleton and those behind the 8th abdominal section for posterior spiracle. The specimens were dehydrated with different concentrations of alcohol 30% to 100%. After being transparented with dimethylbenzene, the specimens were observed and photographed. Under microbioscope the specimens used for image analyzing were made by removing the skin around anterior and posterior spiracle and then spreading thecephalopyngeal skeleton followed by the procedure alone. The image indexes of different parts of anterior spiracle, posterior spiracle and cephalopharyngeal skeleton measured and analyzed by digital image analyzing system.The data obtained were analyzed using the software the curve were drawn by the Excel.Results:1 Morphological characteristics of anterior spiracle1.1At constant temperature, the anterior spiracle of larvae emerged in the duration of second instars in all files tested with the number of fingers varing from 7 to 10 in Aldrichina grahami and Lucilia sericata and 10 to 14 in Chrysomya megacephala, which ranged in a row.Along with the replacement of the age, the color changed from light yellow to heavy yellow gradually and the pigment tube changed from a thin and long type in aged 2 to a short and rough one in 3 aged gradually .1.2 At a constant temperature, the anterior spiracle chamber area of the larvae of Aldrichina grahami, Chrysomya megacephala and Lucilia sericata were minimum at the early time of the second instars, which increase rapidly to the maximum and then declined rapidly into the plateau. The ratio of length to width of anterior spiracle chamber were the maximum at the time of 2nd instars, which decreased the continuously until the end of the 3rd instars. At 20℃to 28℃,the average optical densities of anterior spiracle chamber of each larvae group showed an increasing trend that continued to the end of 3rd instars. But the length and width ratio of anterior spiracle chamber and average optical density of 3rd instars changed more slowly than those of the 2nd instars.2 Morphological characteristics of posterior spiracle2.1 The posterior spiracle structure of Larvae of Aldrichina grahami, Chrysomya megacephala and Lucilia sericata was 1 cleft and no valve at 1st instars that changed to 1 hole and 2 cleft with open valve ring in the 2nd instars and 1 hole and 3 cleft of in the 3rd instars finally. The area increased gradually and the color changed from light yellow to tan, even black gradually.2.2 Average optical densities of posterior spiracle and anterior spiracle chamber of larvae Aldrichina grahami, Chrysomya megacephala and Lucilia sericata changed consistently, Which increased gradually with the rising of time until the end of 3rd instars. These parameters changed more rapidly at 20℃to 28℃in each group during the larval stage and slow down between age periods. Therefore, the average optical density of posterior spiracle is more meaningful than the average optical density of anterior spiracle chamber to determine the age of larvae.3 Morphological characteristics of cephalopharyngeal skeleton3.1 At the constant temperature condition, the color of cephalopharyngeal skeleton tend to become darker, from light yellow to brown and even black with the developing of larvae. The degree and range of chitinazation in cephalopharyngeal skeleton increased in accordance with the development.3.2 At the constant temperature, the area of mouth hook increased periodically. The changes were usually more significant in the period of instars replacing. Once got the maximum, it stopped to increase. The increasing speed of the mouth hook area was faster and the maximum appeared earlier at higher temperatures. The average optical density and the sclerotized area shared the same developing rate. However, these indexes got to the maximal at an later time than the area of mouth hook .3.3 At the constant temperature, the area of larval pharyngeal sclerite also increased periodically. However, the average optical density and the sclerotized area of pharyngeal sclerite increased gradually until the end of third instar. As the temperatures rised, these indexes increased more rapidly and got to the maximal at an earlier time.Conclusions:1 The anterior spiracle chamber area of larvae of Aldrichina grahami, Chrysomya megacephala, Lucilia sericata increased rapidly with time from which was the minimum in the 2nd instars, and once reaching the maximum, decreased rapidly into the plateau. Ratio of length and width of anterior spiracle chamber and average optical density changed in a single trend in the entire process of larval development, keeping an upward or downward trend.2 The average optical density of posterior spiracle and average optical density of anterior spiracle chamber were consistent with the rising of time, which increased of larvae Aldrichina grahami, Chrysomya megacephala and Lucilia sericata to 3rd instars. However, the indicators of growth rate within the same age stage were higher. Therefore, the average optical density of posterior spiracle seemed more important than the average optical density of anterior spiracle chamber in determing the age of larvae. 3 Among the morphology indexes, the indexs of mouth hook kept growing periodically, but keeped steady once reached the maximum in the 3rd instars.4 The average optical density and the sclerotized area of pharyngeal sclerite grew gradually and the increasing speed was kept to the end of third instar.