Correlation of hemocyte counts with different developmental parameters during the last larval instar of the tobacco hornworm, Manduca sexta

Abstract

We determined the changes in hemocyte titer and in the abundance of hemocyte types of the tobacco hornworm Manduca sexta during the fourth and fifth larval stadium and the beginning of the pupal stadium. As we analyzed the samples of individual insects at daily intervals, we were able to correlate phenotypical features, body weight, as well as total protein content and lysozyme activity in the hemolymph with the observations on hemocytes. In the course of the fifth larval stadium, the hemocyte titer decreased slightly and declined further after pupation. Using calculated values for total hemocyte numbers, females had about five times and males three times more hemocytes in the circulating population at the beginning of the wandering stage (in the middle of the fifth larval stadium) than immediately after the last larval--larval molt (from the fourth to the fifth larval stadium). This sexual difference was mainly due to an increase in the number of plasmatocytes, which was more prominent in females than in males. Granular cells were dominant in early fifth larval stadium while plasmatocytes were the most abundant cells in pupae. Oenocytoids and spherule cells disappeared during the wandering stage. Lysozyme activity in the hemolymph rose to a maximum during the wandering stage, with females having lysozyme values twice as high as those for males. These changes in lysozyme activity, however, did not correlate with the increase of total hemolymph protein titer which occurred already at the beginning of the wandering stage. We postulate that changes in hemocyte titers are under direct hormonal control, which has to be proven in future experiments.

Reconfiguration of the immune system network during food limitation in the caterpillarManduca sexta

“…The values found in this study are within the range of those found in earlier studies on food-limited M. sexta (Kramer et al, 1978;Siegert, 1986Siegert, , 1987Siegert et al, 1993;Bedoyan et al, 1992;Ismail and Matsumura, 1992;MeyerFernandes et al, 2001;Beetz et al, 2008). Surprisingly, the caterpillars on the low-nutrition diet managed to maintain their mass, including their dry mass, despite the decline in lipid stores.…”

“…Because immune function in M. sexta changes with age (Eleftherianos et al, 2008;Beetz et al, 2008;Booth et al, 2015), we included a fourth group of caterpillars that were fed on the highnutrition diet for a single day. We wanted to exclude the possibility that the results from the food-limited animals could be explained by developmental delay.…”

A Simple Protocol for Extracting Hemocytes from Wild Caterpillars

“…Methods for extracting hemocytes from medically important insects and lepidopteran model insects have previously been reported 9,14 . Hemocyte extraction methods are adapted according to the insect species, the developmental stage of the insect and its morphological features.…”

The parasitic wasp Cotesia congregata uses multiple mechanisms to control host (Manduca sexta) behaviour

“…Fat body was collected 1 day later (5th instar-day 2) and 3 days later (5th instar-day 4). Because immune function changes with age in M. sexta (Beetz et al, 2008;Booth et al, 2015), we also collected fat body from control 5th instar-day 4 caterpillars to test whether any observed changes in gene expression in 5th instar-day 4 caterpillars could be attributed to ageing.…”