ERRATA. Page 12, lines 16 and 17, for one hundred read three hundred and for one thousand read six hundred. Page 17, line 2, dele first letter in the line. Page 168, line 12, page 177, lines 13 and 14, and page 271, line 10, for Lemna tnsulca read Spirodela polyrMza. Page 209, line 2 of foot-note, after hut insert represents. Page 2;'56, line 7, and page 266, line 19: snoioi n. s. has been shown to be hieroghjijhka, (^. Page 257, insert as line 8 as follows: -ken to the office produced young in ten days. The Page 272, line 13, for P. higuUatus read Pompiliis higuttatus. Page 278, Plate Y., 16, after view insert as follows: a, mentum; 6, labial rudiment; c, maxillary palpi; d, maxilla; e, labrum; /, antenna; g, eye; h, mandible. Page 286, line 11, drop initial the one line. Page 386, line 1, for Comstocki read Comstock. Page 399, line 17, for specimens read specimen. Page 411, line 10, for Michaelson read Michaelsen. Page 441, line 3 from bottom, for 66 read 68. Page 445, line 10 from bottom, for 57 read 5S. Page 466, line 1 from bottom, for Cyima read Oypris. BULLETIN OF THE Illinois State Laboratory OF Natural History. VOLUME IV. Article I. — Bacteria Normal to Digestive Organs of He- miptera. By S. A. Forbes. In 1833 Leon Dufour described and figured in his "i?e- cherches sur les H^miptdres,''* under the non-committal name of "cordons valvuleiix," some curious appendages of the alimentary canal in Scutelleridae, PentatomidaB, and certain Coreidse, misinterpreting their structure, how- ever, and expressing no opinion as to their function, which, in fact, he called a mystery .f These organs had been previously distiuguished by Ramdohr (1811), and they have since been several times referred to by ento- mologists as glands or follicles secreting a digestive fluid, presumably pancreatic. My own entomological studies did not make me particularly acquainted with structures of this class until 1888, in the autumn of which year, while studying the contagious diseases of the chinch bug {Blissus leucoptei'us) I made some dissections of that insect, isolating the alimentary canal and searching dif- ferent parts of it for the source of a bacterial infection discovered in the fluids of chinch bugs when crushed under the microscope. This bacterial development I thus traced to certain coecal appendages of the small intestines so unlike the ^'cordons valvuleux'^ of Dufour's descriptions, that it did not for a time occur to me to connect the two; but in the course of some general dissections of * Pp. 14;)-15l. etc . and figa. 1, 2. 6. 13. 19. 21. + Vp. 150. 171. 2 1111)10Is State Laboratory of Natural TI!story. Heraiptera made at my office this summer by my as- sistant, Mr. John Marten, and myself, it becarne evident that these dissimilar organs could be connected by in- termediate gradations, that they had substantially the same anatomical relations and histological structure, and that all were alike, wherever they occurred, in the highly remarkable fact that they were invariably loaded with myriads of bacteria, differing in genus and species in the different insects, but always confined to these organs.* We further observed that in Coreidae and Lygseidse these coecal structures might be present in one genus and absent in another of the same family, only the higher Hemiptera (Pentatomidse, Scutelleridae, CorimelsenidEe, etc.) invariably possessing them, and the lower Hemiptera invariably wanting them. In case they were absent, their bacterial relationship was never assumed, in whole or in part, by any other organ. The occurrence of "masses of motile vibrio-like objects" in these glands in a Pentatoma was noticed by Leydig in 1857, but I have found no other mention of the matter than that on page 337 of his Lehrhuch der Ilistologie. There are certain coecal appendages of the alimentary canal of other orders of insects, which have a genera^ resemblance to these in Hemiptera, but can be consid- ered homologous with them only in a very loose sense of the word, since they are certainly not homogenous. These gastric pouches in grasshoppers, cockroaches, and carabid beetles do not commonly contain bacteria so far as we have been able to determine. In fact, the only other insect structures in which we have found bacteria normally present with any constancy, were the fatty bodies of various species of cockroaches. It is not ab- solutely certain that these objects from cockroaches are bacteria, as they have not yet been cultivated, our own recent efforts having failed, as did Blochmann's.t * No hibernating specimens have as yet been examined, and it is possible that this phenomenon will be found to disappear with the functional quies- cence of these glands. + Biologisches Centralblatt, Vol. VIL. p. 606. Bacteria Normal to Digestive Organs of Hemiptera. 3 Balbiani's observations, however, (reported in Compter Bendus, Vol, 103, p. 952) to the effect that bacterial forms introduced in the blood of insects are taken up by the cells of the pericardial tissue and destroyed therein, give a certain probability to the hypothesis that these seemino; bacteria of cockroaches are really such. It is true that Balbiani's statements are limited to the peri- cardial tissue in the vicinity of the heart; but as Kowa- levsky has shown* that this tissue is intermingled in many insects with the so-called fatty bodies, it is not unlikely that a more general and critical search would have shown the cells in question to have the same func- tions wherever found. There can, however, be no doubt as to the nature of the objects found in the coecal appendages of the Hemip- tera above mentioned. They not only present every visible characteristic of micrococci and bacilli, but by their leaction to stains, their resistance to prolonged treatment with solutions of caustic potash, and espe- cially and conclusively by the success of our culture ex- periments with both fluid and solid media, they answer to all the tests applicable to the recognition of bacteria. These ccecal structures are probably shown in their simplest form in Pyrrhocoridse (see Dufour, p. 171, and figures 17 and 21), although in the absence of speci- mens of this family for microscopic examination I can only repeat Dufour's surmise that the small and variable ccecal pouches of the small intestine in these Hemiptera are homologues of the complicated apparatus of Anasa and EuHchistus. The next simplest form of this organ which I have thus far seen, is that of the chinch bug) where it consists of five to eight large coeca radiating from a common point of attachment on the intestine about .2 mm. behind the third stomach. These coeca are about .12 mm. in diameter, and average 1.5 mm. in length. They are straight or slightly contorted, with smoothly rounded ends, and are nearly filled, when in normal condition, with large, pale, loosely-attached, sub- • Biologisches Centralblatt, Vol, IX., p. U. 4 Illinois State Laboratory of Natural History. spherical cells, similar to those of the gastric epithelium, and like them usually binucleate, but containing more fatty granules. These cells are varia,ble in size, and in- dividual ones become greatly swollen, and probably break down in secretion. The lumen of the tube is an irregu- lar linear space, not always readily distinguishable in the midst of the cells. That these tubules actually com- municate with the intestine at the point of their inser- tion, I have repeatedly demonstrated under the micro- scope by carefully readjusted pressure on the cover-glass. By this means granules may be made to pass freely from any one of the coeca into the intestine, and even from the third stomach into the coeca through the slender portion of the intestine connecting them. The microbe of these cceca (in the chinch bug commonly Micrococcus insectorum only) occurs primarily in the inter- cellular fluids of these structures, and was excessively abundant in every one of a great number of specimens, the cceca of which I examined separately. My specimens were from all parts of the State of Illinois and from Kansas, and were of various ages, from young immedi- ately following the first moult to the adult. A thorough ^exploration and examination of all the other organs of these chinch bugs failed 1 o discover any trace of this or any other bacterium, with the exception of an occasional infection of that part of the intestine into which the coeca open. Examples of five other genera of Lygseidse have thus iar been dissected by us, in three of which {Lygceus tur- oicus^ Nysius angustatus, and Geocoi*is uUginosus) there is no trace of these "pancreatic" organs, while in two oth- ers {Trapezonotus nebulosus^ and Myodocha senv'j^es) tliej' are present in a stage of development quite above that of the chinch bug, but far below that characteristic of the higher Hemiptera. In Myodocha, for example, they a.re made up of numerous coecal tubes arranged side by side in a single layer, in flat, leaf-like lobes, three in suc- cession, the largest leaf anterior, and the middle one of the series the smallest, the three being bunched together, Bacteria Normal to Digestive OrgOfns of Hemiptera. 5 wrapped arouud the stomach, and imbedded in fatty tissue in a way to require careful dissection for their dis- play. The tube-like structures of which these leaflets are composed are thickest distally, and are attached by their narrow ends to the alimentary canal, which in the first lobe is indistiniruishable from the edge of the sheet itself. This anterior sheet is irregularly palmate, the longest of the finger-like coeca measuring 1.1 mm. and the shortest (those most posterior) about .7 mm. The transverse di- ameter of a single coecum at its broadest end is about .07 mm. This lobe is partly folded together, the folds being held in place by branches of a large trachea, which is distributed abundantly to all parts of the structure. The second or smallest lobe is attached to the intestine by a narrow insertion about .1 mm. behind the preced- ing. It is .5 mm. long by about .2 wide. The third lobe, of medium size, is also attached by a narrow insertion to the intestine immediately beyond the preceding. It is quite regularly palmate in form, is sup- plied by a single much-branched trachea, and measures about .7 mm. long by .5 wide. Crushing successively and separately- all the portions of the alimentary canal upon cover-glasses, and treating by the usual methods for the demonstration of bacteria, I found all the preparations quite free from them, with the exception of those from the above-described leaf-like coecal structures ; and in these, and in every part of them, immense numbers of a minute Micrococcus occurred (not M. insectorum) , situated, as in the chinch bug, chiefly be- tween the large spherical cells of which these bodies were principally composed. Several repetitions of this exper- iment with other specimens gave the same result. In Trapezonotus this organ has the same structure and general appearance as in Myodocha serrlpes. Among the Coreidse I have seen it in Awim tristis, Ahjdm piJosidus, and A. eurinus, but have found nothing resembling it in Corizits lateralis. In this family it has a much more considerable extent than in the foregoing,. 6 Illinois State Laboratory of ]S\(tural History. and now takes the form of rows of shoi-t, transverse coe- cal tubules, standing- in general at right angles to the small intestine, but with their inner ends the smaller. They are fused into a continuous layer, and make hy their arrangement a broad plaited border on each side of the intestine for its whole length, from the stomach to a bladder-like expansion into which the Malpighian organs open. A large trachea runs along the intestine, and its branches are very liberally distributed, right and left, to all parts of these gland-like bodies*. The tubules are lined with a single-layered epithelium very different from that of the part of the intestine into which they open. They may be easily' demonstrated, by pressure under the microscope, to open separately into the ali- mentary canal running along between the rows, and the same fact is evident in stained sections. In everj'- case, again, the intercellular substance within these tubules is little more than a mass of bacteria,—micrococci or bacilli, as far as determined. The same may be said of the PentatomidaB and Cori- melgenidae dissected,—Corimelsena, Peribalus, Mormidea, Euschistus, and Hymenarcys,—except that in these fami- lies there are always four rows of the short transverse tubules instead of two. In Capsidse, Nabidse, Reduviidse, and Aradidae—the only other families examined with this matter in mind—we have not found these structures, and Dufour notes their absence in examples of these families, and in Miris, Phymata, Cimex (Acanthia), and the lower Hemiptera generally. In every case where they have occurred in our dissec- tions, we have made exhaustive search for bacteria in other parts of the alimentary canal also, and in the sal- ivary glands, the fatty bodies, etc., and in all these Hemiptera with only negative results. * The abundant tracheal supply of these organs and the minuto distribu- tion ot the tracheal branches—scarcely less abundant than in the fatty bodies, and much more so than in other portions of the alimentary canal—hint at a peculiar function for this so-called pancreatic apparatus. Bacteria Normal to Digestive Organs of Hemiptera. 7 I have no present desire to speculate concerning the meaning- of the bacterial contents of these glands, but limit myself to this preliminary account, and await the completion of the several investigations in which we are now engaged,—upon the distribution of the coeca, their variations, and their relations to the habits of the spe- cies possessing them, and upon the kinds and nature of the bacteria constantly harbored by these interesting appendages of the alimentary canal. Concerning their relations to insect disease, I will only add that in chinch bugs perishing gradually but rapidly, we find them varying considerably in number in the ccecal appendages; and that w^here they are most abundant, the epithelium of these structures is completely disorgan- ized, only the basement membrane remaining, in the form of coecal tubes filled with a pure culture of Micro- coccus insectorum and a little granular debris, the latter apparently the product of the decomposition of the epi- thelium.