Protective immunity of Nile tilapia against Ichthyophthirius multifiliis post-immunization with live theronts and sonicated trophonts

Two immunization trials were conducted to evaluate host protection of Nile tilapia, Oreochromis niloticus against Ichthyophthirius multifiliis (Ich). Immunizations were done with live theronts or sonicated trophonts by bath immersion and intraperitoneal (IP) injection. The immunized fish were challenged with theronts 21 days post-immunization in trial I and 180 days post-immunization in trial II. The serum anti-Ich antibody and cumulative mortalities of tilapia were determined after theront challenge. Serum anti-Ich antibody was significantly higher (P<0.05) in tilapia immunized with live theronts by immersion or IP injection or with sonicated trophonts administered by IP injection than tilapia immunized with sonicated trophonts by immersion, with bovine serum albumin by IP injection, or non-immunized controls. Host protection was acquired in fish immunized with live theronts by immersion or IP injection. Tilapia immunized with sonicated trophonts by IP injection were partially protected with a 57-77% survival in both trials. At 180 days post-immunization, serum antibody titers had declined in immunized fish yet they were still able to survive challenge. The protection appears not to be solely depending on serum antibody response against Ich.     

Effect of immunization of channel catfish with inactivated trophonts on serum and cutaneous antibody titers and survival against Ichthyophthirius multifiliis

Two trials were conducted to determine the effect of immunization of channel catfish with inactivated trophonts on serum and cutaneous antibody titers and survival against Ichthyophthirius multifiliis Fouquet (Ich). In trial I, catfish were immunized intraperitoneally (IP) with: 1) 1% formalin-inactivated trophonts, 2) 3% formalin-inactivated trophonts and 3) freeze-thawed trophonts. Positive and negative control catfish were immunized with live theronts and 5% bovine serum albumin (BSA), respectively. At day 14, 28 and 50 post-immunizations, no statistical difference was noted in serum or cutaneous anti-Ich antibody titers to formalin-inactivated trophonts or freeze-thawed trophonts. The survival of catfish challenged with live theronts ranged from 33.3% to 43.3% for the formalin-inactivated or freeze-thawed trophonts at 50 d post-immunization. The survival of catfish immunized with live theront and BSA was 93.3 and 0%, respectively. In trial II, catfish were IP immunized with sonicated trophonts at doses of 1) 5 trophonts or 10.2 microg protein g(-1) fish, 2) 10 trophonts or 20.4 microg protein g(-1) fish, 3) 20 trophonts or 40.8 microg protein g(-1) fish, and 4) 5% BSA as the control. Fish immunized with 10 or 20 trophonts g(-1) fish showed highest serum (1/210 to 1/480) and cutaneous antibody titers (1/48 to 1/52), respectively, at 22 d post-immunization and survival (63.3-60.0%). The fish immunized with 5 trophonts g(-1) fish had titers of 1/52 and 1/12 for serum and cutaneous antibody and survival of 23.3%. BSA immunized catfish had background titers and a survival of 6.7%. There was a significant correlation between doses of sonicated trophonts used to immunize and catfish survival (correlation coefficient = 0.859, p < 0.01). These results indicate that doses of sonicated trophonts, but not formalin-inactivated or freeze-thawed trophonts provided both serum and cutaneous antibody responses and survival to live trophont challenge.     

Temperature-dependent protection against Ichthyophthirius multifiliis following immunisation of rainbow trout using live theronts

Rainbow trout Oncorhynchus mykiss Walbaum, 1792 fingerlings were vaccinated by intraperitoneal (i.p.) injection using live theronts of the skin parasitic ciliate Ichthyophthirius multifiliis Fouquet, 1876 at 2 temperatures (12 and 20 degrees C), and protection against challenge infections was subsequently evaluated by bath exposure to live theronts. Vaccination conferred a relative protection (evaluated as the decrease in the number of established theronts) at 12 degrees C and almost complete immunity at 20 degrees C. Significantly increased immobilisation titers (using plasma immobilisation of live theronts) were found in immunised fish at Week 2 and 4 post-vaccination. Lysozyme activity of plasma from vaccinated fish increased from Week 1 to 4. Both immobilisation titers and lysozyme activity were significantly higher at 20 degrees C. This study demonstrated that live theronts are good candidates for an antigen source for development of effective vaccines against white spot disease in this fish host, and further indicated that the protection of rainbow trout against I. multifiliis infection is highly temperature dependent and may be associated with both adaptive and innate response mechanisms.     

Evaluation of a cohabitation challenge model in immunization trials for channel catfish Ictalurus punctatus against Ichthyophthirius multifiliis

Calcein marking and cohabitation challenges have not been investigated in fish parasite research. This study evaluated a cohabitation challenge method in immunization trials against Ichthyophthirius multifiliis (Ich) using calcein, a fluorescent dye, to mark channel catfish Ictalurus punctatus (Rafinesque). Fish were marked by calcein immersion at 0, 500, and 1500 mg l(-1), and then challenged with 15 000 theronts fish(-1). No difference was noted in fish infection levels, mortality, and mean days to death (MDD) caused by Ich between unmarked and marked fish or between fish marked with high (1500 mg l(-1)) and low (500 mg l(-1)) concentrations of calcein. After ensuring that calcein marking had no effect on the susceptibility of fish to Ich theronts, 2 immunization trials were conducted to evaluate the cohabitation challenge model using calcein-marked catfish. Fish mortality, relative percent survival (RPS), and MDD were compared between cohabitation-challenged fish and fish challenged by non-cohabitation. No significant difference was observed in RPS for cohabitation-challenged fish and fish challenged by non-cohabitation. A cohabitation challenge can be used as an alternative challenge method in parasite studies, since it closely mimics natural exposure.     

Cross-immunity and antibody responses to different immobilisation serotypes of Ichthyophthirius multifiliis

Vaccination of channel catfish with either of two serotypes of the parasitic ciliate Ichthyophthirius multifiliis conferred protection against challenge infection by either serotype. Fish were vaccinated by intracoelomic injection with live theronts of isolate G5 (serotype D) or isolate G12 (a new serotype), which express different surface immobilisation antigens. Vaccination with live G12 theronts conferred complete protection against subsequent challenge by both serotypes while vaccination with G5 theronts elicited only partial protection against both serotypes. Vaccination with trophont lysates did not protect against challenge infection. Sera from vaccinated fish were tested in immobilisation assays, ELISAs, and Western blots. Serum antibodies recognised only immobilisation antigens of the serotype used for vaccination in immobilisation assays or on Western blots. No antigens common to both serotypes were identified by Western blots. In contrast, serum antibodies bound antigens in cell lysates from both serotypes by ELISA, demonstrating that antibodies recognising both serotypes are produced in response to infection, which presumably confer observed cross-serotype protection.     

Comparison of serum antibody responses and host protection against parasite Ichthyophthirius multifiliis between channel catfish and channel × blue hybrid catfish

There is limited information available on the immune protection of channel catfish Ictalurus punctatus × blue catfish I. furcatus (CB) hybrid against the fish parasite Ichthyophthirius multifiliis (Ich). The objective of this study was to compare serum antibody response and host protection between channel catfish and CB hybrid catfish using a cohabitation model. Channel catfish and CB hybrid catfish were immunized with live theronts by immersion or by IP injection at the dose of 10,000–20,000 theronts per fish in two trials. The fish were then challenged with theronts to compare serum antibody response and protection against the parasite between channel catfish and CB hybrid catfish. The immunized channel catfish and CB hybrid catfish showed a significantly higher (p < 0.05) serum anti-Ich antibody (titer > 1120) compared to non-immunized controls (titer = 0). After being challenged with live theronts, the immunized channel catfish and CB hybrid catfish had none or a low number of the parasites (<50 trophonts per fish) and showed a significantly higher (p < 0.05) survival (90–100%) than non-immunized controls (0%). Overall results indicated that there was no statistical (p > 0.05) difference on serum anti-Ich antibody, parasite infection and fish survival between immunized channel catfish and CB hybrid catfish.     

Temperature- and salinity-dependent development of a Nordic strain of Ichthyophthirius multifiliis from rainbow trout

During the twentieth century evidence was presented which suggested the presence of various strains and races of the parasite Ichthyophthirius multifiliis Fouquet. However, ecological profiles of various parasite isolates from different climatic zones are sparse. Such stringent characterizations of parasite development at defined abiotic conditions could provide valuable criteria for the different races; profile comparison from various localities is one way to differentiate these strains. Baseline investigations were therefore performed on the associations between abiotic factors (temperature/salinity) and the development of theronts in tomocysts of I. multifiliis isolated from rainbow trout in a Danish trout farm. It was shown that tomocyst formation and theront development took place between 5 and 30°C. Development rates and sizes of theronts were clearly affected by temperature: theronts escaped tomocysts already after 16–27 h at 25°C and 30°C, whereas this process took 8–9 days at 5°C. Likewise, theront size decreased steadily from a maximum of 57.4 × 28.6 μm at 5°C to 28.6 × 20.0 μm at 30°C. This size variation was only partly associated with the number of theronts that appeared at different temperatures. The lowest number of theronts escaping from one tomocyst was indeed found at 5–7°C (mean 329–413). At 11.6, 17.0 and 21°C, the highest number of theronts appeared (mean 546–642). However, at 25 and 30°C, the number decreased (458 and 424, respectively). Additional studies on the salinity dependent development of the parasite (at 11.6°C) showed that salinities above 5 p.p.t. totally inhibited development. Even at 5 p.p.t. the developmental time significantly increased and the number of theronts produced from one tomocyst decreased.     

Relations between histopathology and parasitaemias in Oncorhynchus mykiss infected with Cryptobia salmositica, a pathogenic haemoflagellate

Ichthyophthirius multifiliis is a ciliated protozoan parasite that infects the skin and gills of freshwater fish. This report describes the unusual finding of I. multifiliis within the peritoneal cavities of experimentally infected channel catfish Ictalurus punctatus. Twenty catfish fingerlings were exposed to I. multifiliis theronts using a standardized protocol. Five infected fish and 2 control fish were killed at various time points after infection and their tissues examined. Formalin-fixed, paraffin embedded sections were processed for light microscopy and immunohistochemical detection of I. multifiliis immobilization antigen. Trophonts were observed in skin and gill sections of all exposed fish. Parasites were associated with epithelial hyperplasia, focal areas of cellular disruption and necrosis. In addition to these usual sites of infection, individual trophonts were unexpectedly found within the peritoneal cavities of 4 fish. Staining for parasite antigen facilitated their detection within abdominal adipose tissue or adjacent to intestines. This discovery is interesting as it suggests I. multifiliis may be found in tissues other than the skin and gills during the course of a normal infection.     

Ichthyophthirius multifiliis as a potential vector of Edwardsiella ictaluri in channel catfish

There is limited information on whether parasites act as vectors to transmit bacteria in fish. In this trial, we used Ichthyophthirius multifiliis and fluorescent Edwardsiella ictaluri as a model to study the interaction between parasite, bacterium, and fish. The percentage (23-39%) of theronts fluorescing after exposure to E.?ictaluri was significantly higher than control theronts (~?6%) using flow cytometry. Theronts exposed to E.?ictaluri at 4?×?10(7) CFU?mL(-1) showed a higher percentage (~?60%) of fluorescent theronts compared to those (42%) exposed to 4?×?10(3) CFU?mL(-1) at 4?h. All tomonts (100%) carried the bacterium after exposure to E.?ictaluri. Edwardsiella ictaluri survived and replicated during tomont division. Confocal microscopy demonstrated that E.?ictaluri was associated with the tomont surface. Among theronts released from tomonts exposed to E.?ictaluri, 31-66% were observed with attached E.?ictaluri. Sixty percent of fish exposed to theronts treated with 5?×?10(7) E.?ictaluri?mL(-1) were positive for E.?ictaluri at 4?h as determined by qPCR or fluorescent microscopy. Fluorescent E.?ictaluri were observed on trophonts in skin and gill wet mounts of dead fish. This study demonstrated that Ich could vector E.?ictaluri to channel catfish.     

Partial cross protection against Ichthyophthirius multifiliis in Gyrodactylus derjavini immunized rainbow trout.

Partial cross protection against a skin-parasitic ciliate has been recorded in rainbow trout previously immunized with an ectoparasitic platyhelminth. The susceptibility to infection by Ichthyophthirius multifiliis differed significantly between naive and Gyrodactylus derjavini immunized rainbow trout. Fish partly immune to the ectoparasitic monogenean G. derjavini became less infected and experienced lower mortality than naive fish when exposed to I. multifiliis infections. In vitro studies on immobilization of theronts using decomplemented (heat-inactivated) serum from G. derjavini immune or non-immune hosts showed no immobilization. However, untreated serum from both immune and non-immune fish containing intact complement immobilized theronts (titre 128-256). In addition, non-specific priming of the host response with interleukin (IL-1), bacterial lipopolysaccharide (LPS), concanavalin A (Con A) or mannan did confer a partial resistance to I. multifiliis infection. This will suggest that non-specific factors including complement could be partly responsible for the host response against infections with this ciliate.     

Temperature effects on the immature development time of Culex eduardoi Casal & García (Diptera: Culicidae)

The effect of constant temperatures on the development time from first instar to adult emergence was studied in Culex eduardoi Casal & García reared at 7, 10, 15, 20, 25, 30 or 33°C. Data were adjusted to the linear degree-day model and the nonlinear Briére model. According to the linear model, the development time was inversely related to the rearing temperatures between 7°C and 25°C. Maximum mortality (100%) was recorded at temperatures > 30°C. According to the linear model, the development threshold temperature and thermal constant were 5.7°C and 188.8 degree days, respectively. The lower and upper threshold temperatures and the optimum temperature for the nonlinear model were -2.3, 30.0 and 28.1°C, respectively.     

Two year study on the infectivity of Ichthyophthirius multifiliis in channel catfish Ictalurus punctatus

Ichthyophthirius multifiliis Fouquet (Ich) is a fish parasite that causes serious economic loss for aquaculture. A major difficulty in the maintenance of single isolates of Ich for research purposes is the loss of infectivity. After an unknown number of passages or infection cycles the Ich isolate loses its infectivity. This study determined the infectivity of an Ich isolate during 105 infection cycles in channel catfish Ictalurus punctatus over a 2 yr period. The mean percentage of fish infected by Ich, the infection levels and the time to trophont emergence were each compared after 4 cyclic periods: 1-25, 26-60, 61-90 and 91-105 Ich cycles. Results of this study demonstrated that Ich was significantly more infective (p < 0.05) at 1-25 than 26-105 cycles. Channel catfish were infected at a ratio of 1 infected fish to 8 naive fish at 1-25 and 26-60 cycles. A higher infection ratio occurred at 61-90 and 91-105 cycles. Trophont emergence was noted to be significantly longer at 91-105 compared to 1-25 cycles, during 7 and 5 d respectively, at 23.4 +/- 1.1 degrees C. The results of the present study indicate that the infectivity of I. multifiliis started to decrease after 25 infection cycles and was predominant in the single Ich isolate at 61-90 and 91-105 cycles.     

Comparing tolerance of Ichthyophthirius multifiliis and Tetrahymena thermophila for new cryopreservation methods

Ichthyophthirius multifiliis is an obligate protozoan parasite of freshwater fishes that has a complex developmental cycle. It has not been successfully cryopreserved, so management studies are restricted to parasites obtained during outbreaks or perpetuated by passage in live fishes. To overcome this serious limitation, free-swimming I. multifiliis parasites were tested in a cryopreservation protocol routinely used for a related ciliate, Tetrahymena. In this protocol, I. multifiliis theronts retained infectivity for 3 days, although the protocol itself was ultimately lethal. Exposure of I. multifiliis and Tetrahymena thermophila to a battery of media and cryopreservative reagents showed that I. multifiliis was less hardy than T. thermophila and likely had significant biological and cytoskeletal differences. No combination of reagents, media, freezing rates, or dilution media permitted cryopreservation of I. multifiliis parasites that could then undergo development or infect fish. However, a vitrification protocol was formulated using Ficoll, 1,2-propanediol, and N,N-dimethylacetamide from which intact cryopreserved theronts with some motility were recovered. Understanding the effects of these reagents may lead to both a cryopreservation method for I. multifiliis and to improved understanding of the biology of ciliates.     

Histopathological analyses of native silver catfish Rhamdia quelen (Quoy and Gaimard, 1824) immunized against and challenged with live theronts of Ichthyophthirius multifiliis

As an alternative to treating with chemicals, immunization using Ichthyophthirius multifiliis as a vaccine has been studied in fishes that were often affected with white spot diseases also to understand the possible changes to the tissue caused by the vaccine. The focus of this study was the analysis of the influence of immunization via intraperitoneal injection (i.p.) and via immersion bath (im.b.) on the histopathology of R. quelen after being challenged with live theronts of I. multifiliis distributed in: control (non‐immunized and non‐challenged); non‐immunized and challenged with 12,000 theronts/fish; non‐immunized and challenged with 22,000 theronts/fish; immunized and challenged with 12,000 theronts/fish; immunized and challenged with 22,000 theronts/fish. Water quality was measured in each assay, with 300 fingerlings distributed among 15 tanks with 20 fish in each of three replicates. Six days after challenge, samples for histopathological and parasitological analyses were collected. In both i.p. and im.b. fish the prevalence of I. multifillis in the gills was higher in the non‐immunized fish (33.33% and 27.77%, respectively). Melanomacrophages were present in 53% of the samples of i.p. non‐immunized fish. Fish im.b. immunized and challenged showed more atrophied areas in the hepatocytes. Higher numbers of melanomacrophages in the i.p. non‐immunized fish kidneys were observed compared to control. The results showed no difference in the gill lesions of either immunized or non‐immunized fish compared to control. Histological alterations in the organs of silver catfish were considered light, except in the liver that presented significant atrophy and hypertrophy of hepatocytes after immunization via i.p.     

Effects of cyclophosphamide on the immune system and disease resistance of Asian catfish Clarias batrachus

Cyclophosphamide (CYP), a multifunctional alkylating agent is known as a potent immunosuppressor in endotherms. Here, an experiment was conducted in an ectothermic Asian catfish species, Clarias batrachus to investigate its effect on non-specific and specific immunity as well as disease resistance against a common bacterial pathogen challenge. CYP was intraperitoneally injected for 3 consecutive days at a rate of 200 mgkg(-1) body weight. After 72 h post-injection, control and CYP-treated fish were screened for superoxide production through nitroblue tetrazolium (NBT) assay, myeloperoxidase (MPO) activity, packed cell volume (PCV) and total protein, lysozyme, alternative complement activity (ACH50) and natural haemagglutinin titre as a measure of non-specific immunity level as well as disease resistance against Aeromonas hydrophila challenge to vaccinated and unvaccinated fish. To study the effect on specific immunity, CYP was injected thrice at an interval of 7 days after bacterin injection and serum antibody titre was measured by bacterial agglutination titre assay. The results showed a significant (P<0.05) decrease in NBT and MPO activities, and percent survival against A. hydrophila challenge (both in unvaccinated and vaccinated fish) in CYP-treated fish, when compared to control fish. The above results support the immunosuppressive action of CYP in freshwater catfish, C. batrachus. The neutrophil activities, as measured through superoxide production and myeloperoxidase levels, might be important contributors during A. hydrophila infection and that treatment with CYP reduces phagocytic killing power and inhibits resistance against aeromoniasis.     

Effect of temperature on protein synthesis in fish of the Galapagos and Perlas Islands.

1. The temperature dependency of protein synthesis was studied in vivo in five species of Pacific fish collected in the Galapagos and Perlas Islands: batfish (Ogcocephalus darwini), groupers (Epinephelus labriformis), catfish (Netuma platypogan), puffers (Arothron hispidus) and triggerfish (Sufflamen verres). 2. Liver protein synthesis, assayed by a rapid pulse injection technique, showed a moderate temperature dependency (Q10 = 2-3) in the 15-30 degree C range for all species except puffers (Q10 = 10-20). Synthesis was inhibited above 32 degrees C. 3. Protein synthesis in triggerfish was measured by the constant-infusion technique. Synthetic rates (% of tissue protein synthesized per day) at 25 degrees C were 20% for liver, 10% for gill, 1.8% for red muscle and 0.6% for white muscle. Q10 in the 20 degrees-30 degrees C range was 3.0 +/- 0.5 degrees C for all tissues.     

Cellular and humoral factors involved in the response of rainbow trout gills to Ichthyophthirius multifiliis infections: molecular and immunohistochemical studies

The parasitic ciliate Ichthyophthirius multifiliis infecting skin, fins and gills of fish induces a protective immune response in rainbow trout (Oncorhynchus mykiss) surviving the infection and a similar protection can be conferred by i.p. injection of live theronts. A combined molecular and immunohistochemical approach has been used in this work for pinpointing cellular and humoral immune factors in gill tissue involved in the response and indicating interactions between the systemic and local responses. Fish were immunized by intra-peritoneal injection of live I. multifiliis theronts, control fish were injected with PBS and subgroups were treated with the immuno-suppressant hydrocortisone before fish were challenged with live theronts. Significant up-regulations of genes encoding IgM, IgT, C3, SAA, IL-8, IL-22 and IFN-γ were induced by immunization and challenge. Hydrocortisone treatment had a significant down-regulating effect on genes incoding IgT, IgM, CD4, CD8, IFN-γ, IL-8 and IL-22 in all groups. Immunohistochemistry, using monoclonal antibodies to detect cellular markers, demonstrated active involvement of CD8, MHC II, IgT and IgM positive cells in gill tissue. Putative T-cells (CD8 positive cells) were detected in the intraepithelial lymphoid tissue located at the base of gill filaments and in hyperplastic gill tissue but following infection a clear efflux of these cells was detected. MHC II positive cells were distributed across the gill filaments and accumulated in hyperplastic tissue but hydrocortisone treatment affected their density negatively in both immunized and non-immunized fish. IgT positive cells were present in the epithelial lining of the gill lamellae (suggesting a primary role of this protein in the mucosal defence against the ciliate) whereas IgM positive cells were found only in gill arterioles and the lamellar capillaries. The present work indicates an intensive activity and specialized function of immune cells (B-cells, T-cells and macrophages) and humoral elements such as immunoglobulins IgT and IgM which are orchestrated by cytokines in gill tissue reacting against I. multifiliis.     

Temperature-dependent development and life table parameters of Octodonta nipae (Coleoptera: Chrysomelidae)

The effect of temperature on the development, survivorship, fecundity, and life table parameters of Octodonta nipae (Maulik) (Coleoptera: Chrysomelidae), was studied at seven constant temperatures of 17.5, 20, 22.5, 25, 27.5, 30, and 32.5°C. Preliminary experiments showed that no development was observed at 15 and 35°C. All individuals completed development and females laid eggs from 20 to 30°C. There was a significant decrease in male and female longevity with increasing temperatures from 20 to 30°C. The longest and shortest longevity were 203.5 and 73.7 d for males, and 178.7 and 57.6 d for females, respectively. Females produced on average 62.7, 88.9, 116.8, 70.0, and 47.3 eggs and the life expectancy for a newborn egg was 171.6, 148.7, 114.9, 89.2, and 94.8 d at 20, 22.5, 25, 27.5 and 30°C, respectively. Life history data were analyzed by using an age-stage, two-sex life table. The intrinsic rate of increase (r) and the finite rate of increase (λ) of O. nipae increased with increasing temperatures from 20 to 30°C, while the mean generation time (T) decreased within this temperature range. The r was 0.0155, 0.0249, 0.0339, 0.0361, and 0.0383 d(-1) at 20, 22.5, 25, 27.5, and 30°C, respectively. The net reproductive rate (r(0)) was highest at 25°C (35.0 offspring), and lowest at 20°C (17.0 offspring). T was shortest at 30°C (76.4 d). The results showed that temperature greatly affected the fecundity and life table parameters of O. nipae, and a suitable temperature for population development and fecundity was at 25°C. The life table data can be used for the projection of population growth and evaluation of control programs.     

Effect of temperature on pupa development and sexual maturity of laboratory Anastrepha obliqua adults

The effect of four temperatures (18, 20, 25 and 30°C) on pupa development and sexual maturity of Anastrepha obliqua adults was investigated under laboratory conditions. The results showed that the duration of the pupal stage decreased with an increase in temperature (29, 25, 13 and 12 days, respectively), and maintaining the pupae at 18°C and 20°C results in a low percentage of pupation, pupa weight loss and lesser flying ability. However, it significantly favored sexual behavior, a higher proportion of sexual calls and matings. While enhanced pupa development was observed at a temperature of 30°C, adults had low sexual efficiency, as well as a lower proportion of calls and matings. Gas chromatography-mass spectrometry (GC-MS) analysis of male volatiles showed that the amount of (Z,E)-α-farnesene did not vary among males from pupae reared at different temperatures; however, less (E,E)-α-farnesene was emitted by males obtain from pupa reared at 30°C. Male flies kept at 30°C during their larval stage had more (Z)-3-nonenol and, also, an unknown compound was detected. The fecundity of the females was higher at low temperatures. Regarding fertility, no significant differences were found between temperatures. The optimal temperature on pupa development was 25°C when males displayed ideal attributes for rearing purposes.