The puparium of Cyphomyia wiedemanni Gerstaecker, 1857 (Diptera: Stratiomyidae) found in cassava ( Manihot esculenta ) roots

. The puparium of Cyphomyia wiedemanni Gerstaecker, 1857 is hereby described for the first time, based on 14 puparia from specimens collected buried in soil and feeding off the roots of cassava ( Manihot esculenta Crantz) plants in an area of industrial cassava plantation in Distrito Federal (15°35 ′ 30 ″ S, 47°42 ′ 30 ″ W), Brazil and, reared in controlled lab conditions until the emergence of adults. Unprecedented data on the occurrence of Cyphomyia wiedemanni in Distrito Federal (new record), and the relationship of the larvae with the cassava plants are discussed.

Cyphomyia wiedemanni Gerstaecker, 1857 is widely distributed in the Neotropical region (Fig. 1) (Woodley, 2001;Fachin & Assis-Pujol, 2016).Adults can be found on flowers, but are much more often found sitting on sunny leaves.Like most Cyphomyia species, they display a strong sexual dimorphism, with dichoptic females having, generally, a yellow head, and the holoptic males have a dark head (Fig. 2).Here the puparium of C. wiedemanni is described based on 14 puparia and represents the tenth species of the genus with known immature forms (larvae and pupae), and the first associated with cassava roots (Table 1).sava roots (Manihot esculenta Crantz) at an industrial cassava crop of the Empresa Brasileira de Pesquisa Agropecuária (Embrapa) (15°35′30″S, 47°42′30″W).The larvae were reared in semi-controlled conditions (relative humidity 75%-80% and the temperature 22℃-24℃) in Petri dishes with cassava mass for feeding.After pupation, the specimens were transferred to clean Petri dishes until the emergence of the adults.The adults were pinned, and the puparia fixed in microtubes containing a (1:1) solution of 70% alcohol and glycerol.In total, 14 adults of Cyphomyia wiedemanni (6♂ and 8♀) emerged.All specimens are housed in the Coleção Entomológica do Departamento de Zoologia da Universidade de Brasília (DZUB).
Photographs of the puparia and adults were obtained with the Leica M205C© stereomicroscope equipped with the Leica DFC295© digital camera.Measurements, in millimeters (mm), were made with the Leica LAS-V3-8© software.Micrographs of the puparia were obtained with Scanning Electron Microscopy (SEM).The samples were fixed in osmium tetroxide for one hour.They were first washed in distilled water, and afterwards dehydrated in different concentrations of acetone (30%, 50%, 70%, 90% e 100%), for 10 minutes in each concentration.Then, the samples were critical point dried with CO₂, assembled on stubs, sputter coated with gold and examined in a scanning electron microscope JEOL JSM 7001F©.
The terminology adopted for this study follows Rozkošný (1982)

Comments:
The development time of Cyphomyia wiedemanni, from collection until the emergence of the adults, varied from 85 to 123 days.The larvae and puparia of C. wiedemanni are morphologically similar.A few differences are noticeable, all related to different development stages.We were unable to find features on the surface of the body unique to the larvae or puparia.Both larvae and puparia are flattened dorsoventrally (more pronounced in the larvae than the puparia).The dorsal setae 3 (D3) is always present in the larvae, but is hardly ever observable in the puparia, which are minute, delicate and break easily.In the larvae, the abdominal spiracles are observable dorsally in the segments 2-5, while in the puparia the respiratory horns are present, positioned dorsolaterally.
The general shape of the puparia, as well as the chaetotaxy, have proven conservative across the species of Cyphomyia.Even so, C. wiedemanni exhibits some distinct differences in comparison to the other species, such as: (1) presence of only one ocular tubercle immediately posterior to the eye, shared with C. albitarsis and C. leucocephala, but with a slightly less pronounced sulcus between the eyes and tubercles.( 2 In addition to the record from this study for the Distrito Federal, the distribution of Cyphomyia wiedemanni was expanded to all regions of Brazil based on records from iNaturalist (https://www.inaturalist.org/taxa/294692-Cyphomyia-wiedemanni):North (Acre), Northeast (Alagoas,

DISCUSSION
The terrestrial larvae of Stratiomyidae are cryptic and are generally associated with dim and humid locations like soil, leaf litter, rotten trunks, dung and even in animal carcasses, feeding on the microorganisms associated with the decaying organic matter.An exception to these habits is the larvae of the Chiromyzinae brown soldier-flies genera Chiromyza Wiedemann, 1820 and Inopus Walker, 1950, which live in soil and have adaptations like tubercles on the head and cylindrical bodies that allow for the excavation of soil with a degree of efficiency.Beyond that, they also possess well-developed and functional mandibles that are used to cut plant roots for feeding (Pujol-Luz & Vieira, 2000).Some species cause damage to crops, like Inopus rubriceps (Macquart, 1847) and Inopus flavus (James, 1968), considered sugar cane (Saccharum spp.) pests in Australia (Bull, 1976).In Brazil, the larvae of Chiromyza vittata Wiedemann, 1820 feed on the roots of coffee plants (Coffea arabica Linnaeus), in depths that vary from 30 to 40 cm in soil (D'Antonio, 1991;Pujol-Luz & Vieira, 2000).In the other Stratiomyidae, the mouth apparatus is composed of a mandibular-maxillary complex used for the filtering of microorganisms present in different kinds of substrate (McFadden, 1967).
As in the previous study by Pujol-Luz et al. (2023) with Hermetia teevani Curran, 1934, the larvae of C. wiedemanni were collected buried in soil feeding on the roots of cassava.Hermetia teevani and C. wiedemanni belong to distinct subfamilies, Hermetiinae and Clitellariinae respectively, and do not possess any adaptation or morphological modification related to the excavation of soil or the processing of solid material, as is the case for the immature forms of Chiromyzinae.The presence of larvae of C. wiedemanni in the roots of cassava was probably enabled by previous damage caused by other organisms, as found by Pujol-Luz et al. (2023) for H. teevani.Where, the presence of larvae of H. teevani in the roots of cassava would be a consequence of damage caused by the larvae of cassava borer beetles (Eubulus: Coleoptera, Curculionidae).The cassava borer larvae make galleries in the root's interior and allow for the entry of microorganisms that cause the decaying of the roots (Oliveira et al., 2019).This kind of opportunistic relationship has also been recorded for Cyphomyia.McFadden (1967) collected larvae of C. bicarinata feeding on fluids from damaged parts or decaying tissue from cacti (Opuntia sp.), and James (1957) collected larvae of C. pilosissima in fluid found at the base of Agave plants (Agave sp.), in both cases there was no evidence that the larvae had caused the damage to the plants.McFadden & James (1969) collected larvae of C. erecta in a rodent's nest made in a cavity in a cactus (Platyopuntia sp.).As in previous cases, the presence of the larvae associated with the rodent's nest inside a cavity in the cactus, it is likely due to an opportunistic behavior, since the larvae alone could not be capable of causing damage to the cactus.As such, similar to the findings by Pujol-Luz et al. (2023) for H. teevani, C. wiedemanni would be considered a secondary invader, taking advantage of an opportunity to feed on the damaged roots attacked by the cassava borer (Eubulus).

ACKNOWLEDGMENTS:
We thank Eduardo Alano Vieira, Charles Martins de Oliveira and Josefino de Freitas Fialho (Embrapa Cerrados) for allowing the collection of cassava roots used in this study, Karine Brenda Barros Cordeiro for the acquiring of the SEM photographs, Lucas Pessanha Mousinho for the rearing of the collected larvae and Hélio Ricardo da Silva for reviewing the manuscript.To Martin Hauser for their excellent contribution in reviewing the manuscript.
) Chaetotaxy of thoracic segment 1, similar to that of C. albitarsis, with 2 pairs of anterodorsal setae (Ad₁, Ad₂) and 3 pairs of dorsal setae (D₁, D₂, D₃).(3) Chaetotaxy of abdominal segments also like C. albitarsis.(4) Anal slit length diverges significantly among species, being longer in C. wiedemanni, extending for almost the entire length of the 8 th abdominal segment.(5) Semispherical projections with the dorsal setae (D) on the dorsal surface of the 8 th abdominal segment, as in C. leucocephala e C. albitarsis.
AUTHOR'S CONTRIBUTION: JRPL, FSPG contributed to the study conception and design.The original draft of the text and the figures were prepared by MMRC, FCPG, CVAPL and JRPL.All authors contributed with the reviewing and editing of previous versions of the manuscript and have read and approved the final version.CONFLICT OF INTEREST: Authors declare there are no conflicts of interest.FUNDING INFORMATION: The National Council for Scientific and Technological Development (CNPq) for financial support to JRPL (Grant 305451/2023-5 and Project Immature forms of development of soldier-flies (Diptera, Stratiomyidae), Federal District Research Support Foundation (FAPDF) for financial support to JRPL (Grant 00193-00002181/202).The Coordination of Superior Level Staff Improvement (CAPES) and Graduate Program in Zoology at the Biology Institute of the University of Brasilia (PPGZOO-UnB) to MMRC (Grant 88887.910147/2023-00)and FSPG (Grant 88887.921491/2023-00).

Table 1 .
Species of Cyphomyia Wiedemann: instars and development time, natural substrate breeding and geographical distribution.