Rearing bacteria and maggots concurrently: a protocol using Lucilia sericata (Diptera: Calliphoridae) as a model species

Maggot debridement therapy using live Lucilia sericata (Meigen) larvae is an efficient and cost-effective way to treat chronic wounds. The recent increase in studies to assess the antibacterial properties of L. sericata has created a need for a simple, low-cost, and comprehensible rearing and investigative method for researchers with little or no entomological experience. This paper describes and evaluates a reproducible protocol for sterilising and rearing blowfly larvae utilising two sterile artificial diets (blood–yeast agar and pre-prepared blood agar plates) that is suitable for directly investigating the effect of larvae on microbial growth. Using Lucilia sericata as a model, the results show that larval growth on the pre-prepared blood agar diet is detrimental to larval growth and survival, whereas larval growth and survival on the blood–yeast agar diet are comparable to those of larvae raised on porcine liver. This diet is proposed as a standard for blowfly and bacteria interaction studies investigating clinical microbial strains. Developmental data are provided for L. sericata larvae raised on both sterile and nonsterile diets so that researchers can determine the effect of treatment based on the length of time for larvae to reach the required life stage at 25 ± 2 °C. Information on larval ageing (instars at an average of 1, 2, 3 and 4 days), oviposition times (4–5 days after adult emergence) and adult longevity on the diets (102–116 days) is also given.     

A new method for the production of sterile colonies of Lucilia sericata

Maggot debridement therapy (MDT) refers to the use of blowfly larvae to clean or debride an infected wound. Most commonly, larvae of Lucilia sericata (Meigen) (Diptera: Calliphoridae) are used, and are sterilized prior to use to ensure no further bacterial infections are introduced during treatment. Current methods sterilize eggs from laboratory‐reared blowfly colonies, after which sterile early second instar maggots can be provided to hospitals for use in treatment. Maggots not required for treatment are used for colony regeneration, in which sterility is not maintained. The ability to maintain sterility beyond this would allow further research into fly–bacteria interactions and the effects of different bacteria on the blowfly lifecycle. This study aimed to produce a colony of sterile adults, using current egg sterilization practice, but maintaining sterility through to pupation and emergence. The production of a sterile colony allows further research into the impact of bacteria on fly development and survival. Eggs were placed on a sterile food source within autoclaved plant tissue culture containers to allow growth under sterile conditions. Nutrient agar plating of sterilized and non‐sterilized eggs, larvae and adults (post‐emergence), as well as the pupation medium and feed source in nutrient broth confirmed the aerobic sterility of all samples involved. The lifecycle of L. sericata was successfully completed through pupation to emergence with no effects on lifespan or oviposition by the newly emerged, sterile adult colony.     

饲养五种夜蛾科昆虫的一种简易人工饲料

以黄豆粉、酵母粉及麦麸粉等为主要营养成分 ,研制和筛选出了一种既可工厂化饲养斜纹夜蛾幼虫 ,又可大量饲养甜菜夜蛾、银纹夜蛾、粉纹夜蛾和棉铃虫幼虫的简易人工半合成饲料。利用该饲料目前已实现了工厂化饲养斜纹夜蛾幼虫增殖斜纹夜蛾核多角体病毒     

Behavioural and physiological responses of grass grub larvae (Costelytra zealandica) feeding on protease inhibitors

Abstract

Growth of first instar Costelytra zealandica larvae was significantly reduced after 6 weeks when reared on an artificial diet containing 0.3 and 1% soybean trypsin inhibitor (SBTI), 0.1% and 0.3% potato inhibitor II, and 0.3% potato protease inhibitor I and cowpea trypsin inhibitor. Limabean trypsin inhibitor at 1% significantly stimulated growth compared with that on diet with corresponding levels of added casein. A direct relationship between increased free-trypsin activity and decreased larval growth was observed. Sequential measurement of enzyme activity in third instar larvae feeding on SBTI was compared with that of larvae feeding on casein. The increase in enzyme activity was observed after 14 days in larvae feeding on SBTI. Larvae preferred to feed on SBTI-free diet when given a choice between diet containing this inhibitor at 0.3% and added casein at 0.3%.     

Intestinal bacteria affect growth of Bacillus thuringiensis in larvae of the oriental tea tortrix, Homona magnanima diakonoff (Lepidoptera: tortricidae)

Spores and parasporal crystals of a Bacillus thuringiensis serovar aizawai were fed to fifth instar larvae of the oriental tea tortrix, Homona magnanima, that had been reared aseptically or that had been reared normally. Viable cell numbers of B. thuringiensis and other bacteria in H. magnanima larvae were estimated by homogenization of samples and dilution plating on peptone-polymyxin agar medium for B. thuringiensis cells and on nutrient agar medium for the other bacterial cells. B. thuringiensis did not grow in the larval cadavers of normally reared H. magnanima while bacteria other than B. thuringiensis grew rapidly. In contrast, B. thuringiensis within the larval cadavers of aseptically reared H. magnanima grew and increased 20 times. The bacteria other than B. thuringiensis from the sample homogenates of normally reared larvae that were fed on B. thuringiensis-treated diets had the same characteristics as the bacteria isolated from the guts of healthy H. magnanima larvae, which were putatively identified as Streptococcus spp. and Staphylococcus spp., typical intestinal bacteria of insects. The results strongly suggest that intestinal bacteria influence the growth of B. thuringiensis in the larvae.     

Nutrient balance in medfly, Ceratitis capitata, larval diets affects the ability of the developing insect to incorporate lipid and protein reserves

The Mediterranean fruit fly [Ceratitis capitata Wiedemann (Diptera: Tephritidae)], or medfly, is mass produced in many facilities throughout the world to supply sterile flies for sterile insect technique programs. Production of sterile males requires large amounts of larval and adult diets. Larval diets comprise the largest economic burdens in the mass production of sterile flies, and are one of the main areas where production costs could be reduced without affecting quality and efficacy. The present study investigated the effect of manipulating diet constituents on larval development and performance. Medfly larvae were reared on diets differing in the proportions of brewer's yeast and sucrose. We studied the effect of such diets on the ability of pupating larvae to accumulate protein and lipids, and on other developmental indicators. Except for diets with a very low proportion of brewer's yeast (e.g., 4%), pupation and adult emergence rates were in general high and satisfactory. The ability of pupating larvae to accumulate lipid reserves and proteins was significantly affected by the sucrose and yeast in the diet, and by the proportion of protein to carbohydrates (P/C). In contrast to previous nutritional studies conducted with other insects, low P/C in medfly larval diets (with excess dietary carbohydrates) resulted in pupating medfly larvae having a relatively reduced load of lipids; medfly larvae protein contents in these diets were, as expected, relatively low. Similarly, high P/C ratios in the diet produced larvae with high protein and lipid contents. Differences with other insects may be due to differential post‐ingestion regulation where a high dietary carbohydrate diet reduces the lipogenic activity of the larvae, and induces a shift from lipid to glucose oxidation. Larvae reared on low P/C diets spent more time foraging in the diet than larvae maintained on a high P/C diet, suggesting a compensatory mechanism to complement nutrient intake. The results suggest that the content of brewer's yeast, the most expensive diet component, could be fine‐tuned without apparently affecting fly quality.     

A simple larval diet for population studies on the blowfly Lucilia sericata (Diptera: Calliphoridae)

1. A simple artificial diet was devised for larvae of Lucilia sericata Meigen. 2. A basic diet of 20 g/l agar with 20% horse blood and added yeast sustained normal growth and development, as compared with a lamb meat control. 3. When yeast was added to the blood agar diet at 50 g/l, both peak and final larval weights were increased by 25-50% at higher larval densities (2-8 larvae/g diet) and the hatchling-adult development period was reduced by about 3 days. 4. No adult insects emerged on an agar-yeast diet without blood. Increasing the concentration of blood from 10% to 20% increased adult weight at emergence by 50%, though developmental period was not significantly affected. 5. Increasing larval density significantly reduced the weights of pupae and emergent adults (by up to 50%), both on the blood-agar-yeast diet and the lamb meat control. Percentage survival fell from 80-90% to below 10% on the blood-agar-yeast diet at the highest densities of 4 or 8 larvae/g diet, though survival on lamb was not significantly affected by density.     

Cool-textured diets for use in baculovirus production

The incorporation of baculovirus into semi-synthetic diets has been used for the inoculation of large batches of insect larvae reared under gregarious conditions. The preparation of most artificial diets involves heating to dissolve the agar gelling agent followed by cooling to ～55–60°C whereupon virus occlusion bodies (OBs) are incorporated into the diet. To avoid exposure of thermolabile OBs to high temperatures, a cool-textured diet using soy fiber was evaluated as a simple method to inoculate batches of larvae with the baculovirus OBs. Spodoptera exigua larvae that fed on a diet with 120, 140 or 160?g/l of soy fiber, as a substitute for agar, had a similar weight gain over a 24?h period than larvae that consumed a standard agar diet. The larval weight gain with cool-textured diets with 140?g/l of soy fiber was not improved by the addition of chlorophyll, green food dye colour, or by the removal of ascorbic acid, sodium benzoate and potassium sorbate. Preliminary tests performed to determine the optimal OBs concentration needed for the inoculation of groups of four instar larvae in agar and soy fiber diets revealed no virus inactivation in heated diets with a similar optimal concentration of 36?OBs?mg^?1 for both diets. Moreover, no significant differences were observed in OB production when each type of diet was used to inoculate groups of 250 larvae reared gregariously in plastic containers. Cool-textured diets could be of interest for the production of baculoviruses that require a diet that can be prepared quickly and easily.     

食物种类对淡色库蚊幼虫生长发育和药剂敏感性的影响

淡色库蚊Culexpipienspallens是我国的一种常见家蚊 ,它传播流行性乙型脑炎 ,影响人类健康。研究淡色库蚊生长发育和药剂敏感性与食物的关系 ,对改善蚊虫饲养方法与监测蚊虫药剂敏感性具有重要意义。作者研究了不同食物饲养对淡色库蚊幼虫生长发育和药剂敏感性的影响。结果表明 ,与用面粉 +酵母粉饲养的相比 ,用鼠饲料饲养的幼虫 ,生长发育速度平均要快 2d ,对氰戊菊酯的敏感性则无明显差异。说明可用鼠饲料来代替面粉 +酵母粉饲养淡色库蚊幼虫     

The effect of substrate rearing on growth,aerobic scope and physiology of larval white sturgeon Acipenser transmontanus

The effect of substratum on growth and metabolic rate was assessed in larval white sturgeon Acipenser transmontanus. Yolk‐sac larvae (YSL) were reared in bare tanks or tanks with gravel as substratum from hatch until approximately 16 days post hatch (dph). The effect of an artificial substratum was also evaluated on growth alone. Substratum had a significant effect on mass, with larvae reared in gravel and artificial substrata being larger than those reared without substratum. Routine metabolic rates were significantly lower and relative aerobic scope (the difference between maximum and routine metabolic rate) was significantly higher for YSL and feeding larvae (FL) reared in gravel relative to those reared in bare tanks, particularly before fish started feeding exogenously. Furthermore, gravel‐reared larvae had higher whole‐body glycogen concentrations relative to bare‐tank‐reared larvae. Routine factorial scope (maximum metabolic rate divided by routine metabolic rate) was relatively low in all treatments (< 1·7) indicating a limited ability to elevate metabolic rate above routine early in development and mass exponents for metabolic rate exceeded 1. Taken together, these data indicate that YSL reared without substratum may divert more of their energy to non‐growth related processes impairing growth. This finding underscores the importance of adequate rearing substratum for growth of A. transmontanus and may provide support for habitat restoration and alternative hatchery rearing methods associated with sturgeon conservation.     

Pre-release feeding on yeast hydrolysate enhances sexual competitiveness of sterile male Queensland fruit flies in field cages

Recent laboratory studies of mass‐reared flies in small cages have found that periods of just 24‐ or 48‐h access to yeast hydrolysate can substantially enhance mating performance of mass‐reared male Queensland fruit flies, Bactrocera tryoni (Froggatt) (Diptera: Tephritidae) (‘Q‐flies’). Using field cage tests that provide a better approximation of nature, we here investigated whether access to yeast hydrolysate for 48 h after adult emergence improves the ability of male and female mass‐reared, sterile Q‐flies to compete sexually with wild‐type flies that had been provided continuous access to yeast hydrolysate. Mating probability of sterile males was significantly increased by 48‐h access to yeast hydrolysate; sterile males provided 48‐h access to yeast hydrolysate had mating probability similar to that of wild males provided continuous access to yeast hydrolysate, whereas sterile males deprived of access to yeast hydrolysate had much lower mating probability. Unlike males, access to yeast hydrolysate for 48 h did not increase mating probability of sterile female Q‐flies. We instead found that wild females provided continuous access to yeast hydrolysate had higher mating probability than sterile females that did or did not have 48‐h access to yeast hydrolysate. This result raises the possibility that a bisexual Q‐fly strain might operate essentially as a male‐only release when the flies are given access to yeast hydrolysate during a 48‐h pre‐release holding period. Sterile males given access to yeast hydrolysate for 48 h mated significantly earlier in the evening than wild males and, as in other recent studies, this tendency was associated with an increased tendency to mate on the trees rather than the cage walls. There was no evidence of sexual isolation in this study, as wild and sterile mass‐reared flies showed no evidence of preferential mating with their own kind. Further studies are now needed to assess the potential for pre‐release access to yeast hydrolysate to improve sexual performance and longevity of sterile, mass‐reared, Q‐flies in the field.     

In vitro rearing of the screwworm fly Wohlfahrtia magnifica

Wohlfahrtia magnifica (Schiner) (Diptera: Sarcophagidae) is the main agent of traumatic myiasis in many European, African and Asian countries. Although it can be reared in vivo without technical difficulty, such rearing presents ethical problems. Studies were therefore made of in vitro rearing to facilitate development of laboratory colonies that could be used in a wide range of biological, physiological and applied studies of W. magnifica, particularly in the long period of the year when natural populations of the fly are unavailable for study. Parental colonies of W. magnifica were established from larvae collected from natural infestations of sheep and cattle in central Hungary. First stage larvae were harvested from gravid females and were reared in groups of 5-20 on one of six artificial diets. The diets were based on various combinations of five to seven of eight ingredients: water, agar, blood (heparinized or dried), ground meat, egg yolk, low-fat milk powder, yeast and 10% formol. The larvae were incubated on the diets at 37 degrees C. There was no mortality of first stage larvae, which appeared to feed together in foci, in a natural manner. However, during the second stage, and especially after renewal of diet associated with disturbance of the larvae, many larvae began to disperse, crawling over the surface of the media and feeding less intensively. Mortality of larvae during all larval stadia was 64-98%, compared to 33% in batches of third stage larvae collected from natural infestations. The mean weights of puparia from artificial diets ranged from 38.7 to 59.3 mg, compared to 92.2 mg of puparia from larvae collected from natural infestations. There was a high mortality in the pupal stage, from 61 to 100%. Only a maximum of 6% of first stage larvae were successfully reared to the adult stage. Further studies are needed to identify factors present or absent in the diets that contributed to the present poor development of W. magnifica in vitro.     

Temperature influences swimming speed, growth and larval duration in coral reef fish larvae

The effects of temperature on growth, pelagic larval duration (PLD) and maximum swimming speed were compared in the tropical fish marine species Amphiprion melanopus, to determine how temperature change affects these three factors critical to survival in larvae. The effects of rearing temperature (25 and 28 °C) on the length of the larval period and growth were examined in conjunction with the effects of swimming temperature (reared at 25 °C, swum at 25 and 28 °C, reared at 28 °C, swum at 25 and 28 °C) on critical swimming speed (U-crit). Larvae reared at 25 °C had a 25% longer pelagic larval duration (PLD) than larvae reared at 28 °C, 12.3 (±0.3) days compared with 9 (±0.6) days at 25 °C. To offset this effect of reduced developmental rate, growth and U-crit were measured in larvae reared at 28 and 25 °C at the same absolute age (7 days after hatching (dah)) and same developmental age (7 dah at 28 °C cf. 11 dah at 25 °C), corresponding to the day before metamorphosis. Larvae reared at 25 °C were smaller than larvae reared at 28 °C at the same absolute age (7 dah at 25 °C cf. 7 dah at 28 °C), yet larger at similar developmental age (11 dah at 25 °C cf. 7 dah at 28 °C) when weight and standard length were compared. This stage-specific size increase did not result in better performance in larvae at the same developmental age, as there was no difference in U-crit in premetamorphic larvae reared at either temperature (7 dah at 28 °C c.f 11 dah at 25 °C). However, U-crit was considerably slower in 7-day-old larvae reared at 25 °C than larvae of the same absolute age (7 dah) reared at 28 °C. Swimming temperature controls demonstrated that a change in temperature immediately prior to swimming tests did not effect swimming performance for larvae reared at either temperature.A decreased in rearing temperature resulted in longer larval durations, reduced growth rates and slower swimming development in larvae. However, the magnitude of the response of each of these traits varied considerably. As such, larvae reared at the lower temperature were a larger size at metamorphosis but had poorer relative swimming capabilities. This study highlights the importance of measuring a range of ecologically relevant traits in developing larvae to properly characterise their relative condition and performance in response to environmental change.     

Rearing of the colydiid beetle predator, Dastarcus helophoroides, on artificial diet

Artificial diets for the colydiid beetle, Dastarcus helophoroides, a predator of cerambycid beetles and xylocopid bees, were evaluated. Hatched larvae were reared on artificial diets composed of silkworm pupa-powder, dry yeasts, yeast extract, sucrose, peptone, squid liver oil, preservatives and distilled water, but their emergence rate was very low. If larvae were fed paralyzed cerambycid larvae till they became approximately 8 mm in body length and then reared on artificial diet, emergence rates were high. Using the latter method, mass-production of this colydiid beetle is feasible.     

DNA Taxonomy and the identification of immature insect stages: the true larva of Tauriphila argo (Hagen 1869) (Odonata: Anisoptera: Libellulidae)

For many insect taxa, larval morphology plays a decisive role in various fields like taxonomy, phylogeny or ecology. However, species identification is usually based on imaginal characters and the identification of larvae depends upon an established link to unequivocally identified imagines. This taxonomic correspondence of larvae and imagines is far from being established in many odonate species. We have employed a molecular approach to link larval and adult specimens in Tauriphila argo (Hagen, 1869). The sequenced mt SSU gene fragments of the reared female, supposedly a T. argo female, and a clearly identified male specimen of the species were identical. However, the larva of the reared female clearly differed from the described T. argo larva, previously matched to the species. From this observation, we conclude that the previously described larva of T. argo does not belong to this species because of too many phenotypic differences that far exceed the generally observed intraspecific variation.

It can be foreseen that the molecular approach will prove to be effective in identifying unknown larvae in many insect species. Additionally, the discrimination of sibling species or the linkage of allotypes and holotypes will become feasible with this approach.     

Induction of antibacterial activity in larvae of the blowfly Lucilia sericata by an infected environment

Maggot debridement therapy (MDT) is a method for the treatment of intractable, infected and necrotic wounds. In MDT, sterile larvae of Lucilia sericata Meigen (Diptera: Calliphoridae) are applied to infected wounds, where they exert antibacterial effects. Once the larvae are placed in the wound, they are no longer germ-free. This study analysed the influence of infected environments on larval antibacterial activities. Sterile larvae were mixed in a test tube containing a bacterial suspension of Staphylococcus aureus or Pseudomonas aeruginosa, transferred to liver puree agar, and incubated at 25 °C for set periods. To collect the larval extracts, the incubated larvae were transferred to a test tube containing phosphate buffered saline (PBS), cut into multiple pieces with scissors, and centrifuged. The supernatant was used to test antibacterial activities. The results showed that infected larvae had better antibacterial capacities than sterile larvae. Antibacterial activities were induced by pretreatment with a single bacterial species, S. aureus or P. aeruginosa, within 24 h and 12 h, respectively, and disappeared after 36 h. The activities were effective against S. aureus, but not against P. aeruginosa. This natural infection model is very similar to the clinical wound context in MDT and will be a powerful tool with which to study the antibacterial activities of L. sericata larvae in MDT.     

The in vitro development from egg to prepupa of Campoletis sonorensis (Hymenoptera: Ichneumonidae) in an artificial medium: importance of physical factors

A further attempt to improve the in vitro rearing of the solitary larval endoparasitoid Campoletis sonorensis (Hymenoptera: Ichneumonidae) was made after third instars were obtained in previous studies. Respiration was an important factor for the parasitoid's continued development: both small diet volumes that exposed the larval cuticle to air and O(2)greatly improved growth and development of C. sonorensis. Fifth instars were obtained when the parasitoid larvae were reared in medium volumes of 0.25ml and 0.1ml per well. Increasing the O(2) tension of the air resulted in larger larvae and prepupae. The agar layer under the nutritional medium was designed to slowly remove water from the diet. The results indicated that a drier environment was important for the parasitoid at the terminal stage. One percent and 0.8% agar gels, which removed water from the diet more rapidly, provided a drier environment and resulted in improved larval growth compared to the 0.6% agar gel. The detrimental effect of higher concentration of chicken egg yolk mixed in the diet was eliminated by using a dry egg yolk layer covered by an agar layer. This sandwich technique resulted in more parasitoid larvae molting to the final larval instar (5th) and the formation of many prepupae. However, the development of prepupae was premature and none of the fifth instar larvae successfully formed a normal cocoon.     

Digestive enzymes in larval Coregonus lavaretus L.

Using histochemical methods, morphofunctional aspects of the alimentary tract of larval coregonids were investigated. Larvae of Coregonus lavaretus were reared for 34 days with either zooplankton or one of two dry diets. Ontogeny, localization and diet-related modifications of the following enzymes were examined: trypsin (luminal digestion), aminopeptidase, maltase, alkaline phosphatase (brush border-bound digestion) and unspecific esterase (intracellular nutrient processing). All of the enzymes studied were present in 13-day-old larvae. Except for the intracellularly located unspecific esterase, there was an ontogenetic enhancement of enzyme staining intensities accompanied by a significant increase in the volume of the intestinal mucosa. Enzyme activities differed within and between intestinal regions. This finding suggests that a spatial gradient of nutrient breakdown and absorption already exists in the morphologically and physiologically incompletely developed digestive system of larval coregonids. Digestive enzyme activities were modified in response to the dietary regimen. There was no obvious correlation between enzymic response and growth performance of the larvae.     

New larval agar-based diet for laboratory rearing of Mediterranean fruit fly <Emphasis Type="Italic">Ceratitis capitata</Emphasis> (Diptera,Tephritidae)

Five larval diets for laboratory rearing of Ceratitis capitata were tested. These diets were based on wheat bran, microcellulose, potato starch and agar. To evaluate the quality of diet, pupal rearing efficacy and pupal weight were checked. The best results were obtained with an agar based diet used for Manduca sexta laboratory rearing. To simplify the preparation and to reduce the cost of this diet, a new formulation was developed. Larvae reared on the new agar-based diet achieved higher pupal rearing efficacy than larvae reared on bran diet recently used in medfly mass rearing facilities. Heat treated medflies reared on the new agar-based diet achieved similar pupal rearing efficacy with heat treated medflies reared on bran diet. When testing population density, higher pupal rearing efficacy was again achieved on new diet. The highest pupal rearing efficacy was achieved with 100 eggs per 25 g of diet, lowest with 500 eggs per 25 g of diet. Concerning pupal weight, there was no difference in results achieved on Petri dishes with different larval population densities. Larvae reared on new agar-based diet reached better results than larvae reared on bran diet. The preparation of the new diet is simple and the cost is low, so it is good for laboratory tests and rearing.     

Effect of probiotic adult diets on fitness components of sterile male Mediterranean fruit flies (Diptera: Tephritidae) under laboratory and field cage conditions

The aim of the current study was to investigate the effect of probiotic adult diets, i.e., adult diets containing viable symbiotic intestinal bacteria, on the pheromone-calling activity, mating success, life expectancy, and survival of mass-reared male Mediterranean fruit flies, Ceratitis capitata (Wiedemann), as an avenue for improving the field performance of sterile males in release programs to eradicate, suppress, or prevent spread of wild populations. The effect of inoculation of two standard adult diets (sugar-yeast granulate [SY] and sugar agar [s]) and two experimental formulations (yeast-reduced granulate [Sy] and yeast-enhanced sugar agar [sy]) with Enterobacter agglomerans and Klebsiella pneumoniae (typically occurring in the gut of wild flies) on the different fitness components was assessed in the laboratory and on field-caged host trees. We found that, in the laboratory, males reared on the probiotic yeast-enhanced agar, sy, had a significant mating advantage over competitors fed the standard s agar (probiotic and control) or noninoculated sy agar; no effect of probiotic enrichment (or lowering the yeast content) was found with the granular diets. Mating test results obtained in the field were inconsistent with laboratory data in that no differences in the numbers of matings were observed between males reared on any of the probiotic and control agar diets (or the SY granulate), whereas males feeding on the probiotic modified granulate, Sy, scored significantly more matings than their control competitors. The pheromone-calling activity of males maintained on the granular diets was not affected by probiotic enrichment on any of the seven observation days. Agar-fed males, however, "called" more frequently on days 6 and 7 (but not on days 1-5) when their diet contained the probiotic load. Laboratory survival of granulate-fed males was found to be significantly prolonged with probiotic inoculation and lowering the yeast content of the standard SY granulate (but not with probiotic inoculation of sy). Similarly, males reared on the probiotic and control modified agars (sy) survived significantly longer than those feeding on the standard s agars (inoculated and control). Again, the results obtained in the field were inconsistent, because no differences between treated and control males were found for any of the diets. The findings are discussed in the light of other published studies on adult nutrition and behavioral ecology in C. capitata.