Sperm ultrastructure of Pochazia shantungensis (Chou & Lu) and Ricania speculum (Walker) (Hemiptera, Ricaniidae) with phylogenetic implications Author Jiang, Zhen Author Liu, Jianing Author Qin, Daozheng text ZooKeys 2019 880 43 59 http://dx.doi.org/10.3897/zookeys.880.32810 journal article http://dx.doi.org/10.3897/zookeys.880.32810 1313-2970-880-43 EFFE7128E4A242F684CE0BDDD5E0A447 E55FD13BE3335AB19EED10E036E9ABD5 Pochazia shantungensis (Chou & Lu, 1977) Description. The mature spermatozoa of P. shantungensis are held together (totally 128 spermatozoa per spermatodesm) in the form of coiled sperm bundles in the seminal vesicles. Anterior ends of heads are embedded in a homogenous matrix that forms the spermatodesmata ( Fig. 1A, B, D ). The spermatozoon is long and filiform when it is separated from the bundles and exposed to a 0.9% saline solution ( Fig. 1C ). It is approximately 127 µm in mean length and has an elongate single head (about 16 µm ) and a conventional single flagellum (about 111 µm ). Figure 1. TEM and light micrographs of spermatozoa and spermatodesms of P. shantungensis . A, B Light micrographs of spermatodesm and spermatozoa C light micrograph of a single spermatozoon with the head ( H , arrow) and wavy flagellum ( F ) D, E TEM micrographs of cross-sections of spermatozoa, showing the acrosome ( a ) and nucleus ( N ). Arrow shows head cluster, asterisk indicates the acrosome. Scale bars: 20 µm ( A-C ); 2 µm ( D ); 0.5µm ( E ). The head is formed by the nucleus and the acrosome. The acrosome has an irregular saccular acrosomal vesicle and a perforatorium, both located anterior to the nucleus ( Figs 1E , 2A, B , 3A, B ). Between the base of the perforatorium and the anterior portion of the nucleus is a noticeable transition ( Figs 1E , 2B , 3 C-E ). The acrosome gradually invaginates posteriorly to form a subacrosomal space in which the anterior part of the elongated nucleus is inserted ( Figs 2B , 3C, D ). The nuclei, different in shape ( Figs 2B , 3 B-N ), are full of homogeneous condensed chromatin and are separated from each other by a cell membrane ( Fig. 1D ). The ovoid nucleus measures approximately 0.94 µm in diameter in cross-section; in longitudinal-section, it turns into a cylinder-shape ( Fig. 2A, B ). Figure 2. Longitudinal sections of spermatozoa of P. shantungensis . A, B Spermatozoa, showing acrosome ( a ), nucleus ( N ), arrow indicates connection area between acrosome and nucleus C nucleus-flagellum transition, showing nucleus ( N ), mitochondrial derivatives ( md ), accessory body ( ab ), axoneme ( ax ), arrow indicates centriole, triangular arrowhead indicates centriolar adjunct D, E flagella of sperm, showing axoneme ( ax ), mitochondrial derivatives ( md ) and cristae (arrow). Scale bars: 2 µm ( A ); 0.5 µm ( B, D ); 0.1 µm ( C, E ). The nucleus-flagellum transition region has a centriole and centriolar adjunct ( Fig. 2B, C ). The centriole starts near the terminal incurvation of the nucleus and terminates anterior of the axoneme ( Fig. 2B, C ); it is parallel to the moderately electron-dense centriole adjunct ( Fig. 2B ). The centriole adjunct contains dense granules between the accessory bodies and mitochondrial derivatives in longitudinal profile ( Fig. 2B, C ); in cross-section the centriole adjunct arises near the end of the nucleus and terminates anterior to the mitochondrial derivatives ( Fig. 3 I-K ). The nucleus is wrapped by the centriolar adjunct ( Figs 2B, C , 3 I-M ). In several transverse profiles of spermatozoa, the posterior nucleus region overlaps the different regions of accessory bodies and mitochondrial derivatives ( Fig. 3 G-L ). Figure 3. Cross-sections of spermatozoa of P. shantungensis . A Acrosome, showing the dothideoid acrosome B-E serial cross-sections of head showing the dothideoid acrosome ( a ), the nucleus ( N ), and the plasma membrane ( pm ) F oval nucleus ( N ) G-N nucleus-flagellum transition region, showing the nucleus ( N ), mitochondrial derivatives ( md ), accessory bodies ( ab ), axoneme ( ax ). The asterisk indicates the centriolar adjunct ( ca ). Scale bars: 0.5 µm ( B-D ); 0.2 µm ( A, K, L, N ); 0.1 µm ( E-J , M ). The flagellum region contains an axoneme, two mitochondrial derivatives and two accessory bodies ( Fig. 4 A-D ); they are parallel to each other throughout most of the length of the flagellum ( Fig. 2 B-D ). The axoneme arises from the centriole ( Fig. 2C ). It is composed of two innermost microtubules, nine outermost accessory microtubules, and nine doublets, showing the typical 9 + 9 + 2 microtubules arrangement in insects ( Fig. 4D, G ). The mitochondrial derivatives and accessory bodies are symmetrical in size and diameter in cross-section ( Fig. 4 A-D ). Each mitochondrial derivative is made up of one serrated electron-dense area, one small oval electron-lucid portion and one mitochondrial cristae region ( Fig. 4C ). In longitudinal-section, the mitochondrial derivatives are positioned lateral to the axoneme and are initiated near the extreme base of the centriole adjunct ( Fig. 2 B-D ). The cristae are perpendicular to the longitudinal axis, bearing regular intervals (42 nm) between adjacent derivatives ( Fig. 2E ). The accessory bodies are fishhook-shaped; they originate from the centriolar adjunct between the axoneme and mitochondrial derivatives ( Fig. 4 B-D ). Close to the posterior sperm tip, the axoneme becomes disorganized step by step, and the accessory bodies gradually taper to a cone-shape, while the mitochondrial derivatives disappear ( Fig. 4E, F ). At the terminal region of the flagellum, the doublet microtubules are the last to disappear ( Fig. 4H ). Figure 4. Cross-sections of the sperm flagellum of P. shantungensis . A-D Flagella, showing axoneme ( ax ), fishhook-shaped accessory bodies ( ab ), D-shaped mitochondrial derivatives ( md ), containing oval lucent region ( 1 ), serrated electron-dense region ( 2 ) and mitochondrial cristae region ( 3 ) E-F flagellum, mitochondrial derivatives slowly disappear, axonemes ( ax ) become disordered, accessory bodies ( ab ) become smaller G axoneme, showing the typical 9 + 9 + 2 pattern, nine outermost accessory microtubules ( am ), nine doublet microtubules ( dm ) and two innermost central microtubules ( cm ) H Showing doublet microtubules finally disappearing. Scale bars: 2 µm ( A ); 0.5 µm ( B ); 0.1 µm ( C-H ).