Embryo Development Of The Parthenogenetic,Sexual And Hybrid Eggs In The Lower Termites

Termites are known as the oldest social insects worldwide.Presently in China total 473species of termite belonged to 4 families and 44 genera have been recorded,among which 111Reticulitermes species are widely spread in different zones of China.It is such a tough job,to sample and detect the eggs of termite.Because of their cryptic lifestyle and embryonic development,few reports are presented.The dispersion flight season of these Chinese Reticulitermes species are usually started from February to June,but in some regions different species are distributed,sharing their boundaries and having overlapping flight season.These reasons become essential sources of hybridization between two different heterospecific populations of termites.In the present work,Reticulitermes chinensis,R.aculabialis,R.labralis and R.flaviceps were selected for studies on interspecific hybridization,embryo development and morphology of gonads.The main results are as follows.?1?The developmental stages of R.chinensis were observed by using basic techniques of paraffin section,shell softening of fertilized and unfertilized eggs under.The ideal condition of shell softening for fertilized egg is 1 min treatment with an applied solution of 10ml CH3COOH with 10%KOH.While for unfertilized eggs,the solution?glacial acetic acid15 ml combined with 10%KOH?was treated for 1.5 min,that has given the highest rate of softening successfully.?2?Different developmental stages of Reticulitermes embryo were categorized through the study of paraffin sections.The termite eggs after insemination about 2 to 3 h,form a zygote,and then the mitosis began as superficial cleavage.The blastoderm cells near the micropyle were joined to each other and form a dense and uniform band within 24 to 48 h,called blastoderm.The blastoderm formed the proximal area of the embryo at 48 to 96 h,and the blastoderm cells at one side of the ovum become thicken and thicken thus made an embryonic band.The embryonic band is regularly elongated to increase the body parts from the front to back at 3 to 5 d.The germinal layer continues up to develop into a U-shaped morph,a tail twist at 8 to 10 d and a double-bend structure at 10^th to 12^th d.At 12^th to 15^th d,the internal organs of embryo progressively began to develop.Furthermore,at 15^th to 25^thh d,the 1^st rotation and 2^nd was seen around the elongated axis.While,on 25^th to 30^th,the internal organs of the embryo were developing continuously 25 to 30 d.The head of the embryo observed vigorously squeezing the eggshell at 28 to 30 d,and the eggshell at the back squeezed.?3?The??eggs?unfertilized eggs?and??eggs?sexual eggs?were selected to examine the cleavage of R.aculabialis and R.chinensis.The Laser scanning confocal microscope?Olympus,FV1000?and digital microscope?Keyence,VHX-5000?were used for examination of egg cleavage,and shape,volume,width,and length of embryos.In all developmental stages of R.aculabialis embryos in unfertilized and fertilized eggs observed healthy except change in number of nuclei.It was demonstrated that the unfertilized eggs and fertilized eggs of R.aculabialis have the same rate of cleavage.In fertilized eggs of R.chinensis,and R.flaviceps all developmental stages of embryos were healthy.While,the unfertilized eggs of R.chinensis and R.flaviceps cleaved usually,but almost their development was observed abnormal,and at the end they could not hatch.The two types of R.chinensis embryos showed obvious differences on length,width and twist in similar stages of development.The width and length of sexual and fertilized eggs of three termite species changed on the 5 d,10 d,15 d,20 d,25 d,and 30 d at similar phase of development.The developmental studies of the embryonic stages of hybrid eggs were carried out by cytological,histological and morphological observation in the whole incubation period from the zygote cleavage to hatching.All observation of hybrid zygote cleavage,blastodisc formation,blastoderm development and elongation,and germinal layer development into the head,thorax and abdomen appendages revealed that the whole embryogenesis following the systematic developmental pattern of the sexual and parthenogenetic embryo.However,the hybrid development was a more natural,repaid process and earlier than the sexual and parthenogenetic eggs without any anomaly.The earlier hatching was seen in the hybrid eggs such as Ra?×Rc?,Ra?×Rc?,Ra?×Rf?and Rc?×Rf?at 28^th and 29^thh d.Even an increase in number of nuclei,length and width of hybrid eggs during 5 h,24 h,and 48 h observation were contrast to sexual and parthenogenetic eggs.It was determined that the micropyle of one species egg accepted sperm of another species for fertilization without any resistant of a female barrier.Might be the egg fail to identify the sperm of another species.Anyhow the hybrids have more advantages over sexual and parthenogenetic development according to the Darwen theory of natural selection.?4?The R.chinensis,R.aculabialis,R.labralis and R.flaviceps were paired as Rc×Rf,Rl×Rf,and Rc×Rl,respectively for heterospecific hybridizations.Genotypes of offspring and parents of Rf?×Rf?,Rc?×Rf?,and Rc?×Rf?were distinguished by DNA capillary electrophoresis in order to know the occurrence of genetical barriers,to search the probability of hybrid colonies and new species of termite in near regions.For Ra×Rc,between the two heterospecific pairs,the time required for egg-laying,and relatively low rate of the survival ship of delates was observed,they have a tendency to be steady after six months.The number of the eggs at 30^th days from the start of egg laying and offspring after 6^th months were both lesser amount than those of the homogeneous pairs,with significant alteration.In the initial conspecific colony of R.aculabialis delates the rate of death was relatively high contrast to R.chinensis.While the lowest rate of survival was seen in Rf×Rl colonies.Although the swarming season and distribution of Reticulitermes species are altered,that way heterospecific pairings of Reticulitermes termites produce progeny under laboratory conditions,and there were no pre-zygotic barriers in a hybrid.In other colonies,the rate of survival of other colonies was seen the highest,while the lowest rate of survival was seen in Rf×Rl colonies.Although swarming season and distribution are altered,the heterospecific pairing of Reticulitermes termites produce progeny under laboratory conditions,and there were no pre-zygotic barriers in a hybrid.At foundation of conspecific and heterospecific colonies of R.aculabialis,R.chinensis,and R.flaviceps workers,there no chasing of female was observed in any pairing types during tandem pairing.At foundation of conspecific and heterospecific colonies of R.aculabialis,R.chinensis,and R.flaviceps workers,there no chasing of female was observed in any pairing types during tandem pairing.The earliest nest at 1-2 d was excavated by FF,and FM workers.Onward eggs laying were observed early at27 d,32 d,and 33 d in FF pairs of Rf?×Rf?,Rc?×Rc?,and Ra?×Ra?that followed by FM pairs Rf?×Rf?,Ra?×Ra?,Rc?×Rc?at 32 d,34 d and 35 d correspondingly.Similarly,Rf?×Rc?at the 33 d,Rf?×Ra?at 35 d,Rf?×Ra?at 35 d,Ra?×Rf?at 36 d,Rc?×Rf?at36 d,and Ra?×Rc?at 38 d respectively of colonies foundation.Additionally,the number of eggs laid per-month by FF pairs in Rc?×Rc?were 14.20±2.30,Ra?×Ra?pairs 11.33±4.40and Rf?×Rf?7.40±2.17 followed by the FM pairs Rc?×Rc?8.10±2.40,Ra?×Ra?pairs7.00±2.00 and Rf?×Rf?7.00±2.00.Moreover,in hybrid pairs of Rc?×Rf?,Ra?×Rf?,Ra?×Rc?,Rc?×Ra?,Rf?×Ra?and Rf?×Rc?were 13.00±3.55,12.90±4.80,12.20±2.57,12±5.00,7.60±2.20,and 6.00±3.30 respectively.After parthenogenetic and sexual reproduction,hybridization was also discovered one of the most active,energetic,advance and vigorous mean of reproduction of R.aculabialis,R.chinensis and R.flaviceps.?5?The part of this study was conducted based on morphometric analyses and histological examinations of the sperm,spermatogenesis,testes,and spermathecae in two capable hybridized termite species,R.flaviceps and R.chinensis.The accessory glands of males were found without spermatozoa in both Reticulitermes species,while the patterns of spermatogenesis in alate testes of both species were found similar.During spermatogenesis,the gametes?spermatogonia,spermatocytes,spermatids,and sperm?produced in a sequence of essential developmental processes.The aflagellated sperm has been transferred by males numerous times a day after few copulations,and they were stored in spermatheca and used for fertilization.The nutrition was obtained by the sperm from the secretory cells,for a living and being functional during storage in the spermathecae.Physiologically females contributed nutrients to keep the sperm alive and energetics inside their bodies as noticed in semen.The Reticulitermes species have a pear or finger-shaped spermathecae.The spermathecae had thick walls that were divided into four types of cells?epithelial,muscle,secretory and tube cells?.Anatomically spermathecae of both termite species consisted of lumens,ductules,few glands,and small hair-like projection.Hence,R.flaviceps and R.chinensis are two different species,but on the base of morphology,physiology,anatomy,and histology of the sexual organs they are closely related and since they can hybridize under normal conditions.Since Reticulitermes species can hybridize under experimental conditions to produce offspring,the similarity in the structure of sexual organs of heterospecies provides possible evidence for the successful hybridization.