Moniliformis Dubius Research Articles

The Utilization of C 14 O 2 in the Production of Acid Metabolites by Moniliformis dubius (Acanthocephala)

The fixation of carbon dioxide (NaHCO0) by Moniliformis has been studied. This parasite, incubated in NaHC403 for periods ranging from 30 sec to 5 min, incorporated label into aspartic acid, alanine, serine, malate, succinate, fumarate, and lactate. Pyruvate, oxaloacetate, and a-ketoglutarate, identified as 2,4-dinitrophenylhydrazine derivatives, were found but only the two former compounds were labeled. The pattern of labeling indicated that fixation into pyruvate or phosphoenolpyruvate had occurred. Moniliformis dubius was found to produce no carbon dioxide during anaerobic metabolism (Laurie, 1957). Acids excreted during aerobic metabolism were tentatively identified as acetic, formic, and lactic acids (Laurie, 1959). The present author studied the distribution of label in organic and amino acids in M. dubius incubated in glucose-U-C14 (Graff, 1964). The pattern of labeling indicated that carbon dioxide fixation had occurred at a significant level. The above observations initiated the present study in which M. dubius was exposed to NaHC1403. MATERIALS AND METHODS Female worms were removed, after 42 days of infection, from male Sprague-Dawley rats, which were reared and infected as previously described (Graff and Allen, 1963). Approximately 1 g of female M. dubius was weighed in each case and preincubated for 15 min in buffered mammalian Ringer's solution [pH 7.5 with 0.03 M tris (hydroxymethyl) aminomethane]. Worms were removed, blotted, and transferred to flasks containing 5 ml buffered Ringer's and 10.0 gM NaHC03 (sp. activity 1.89 mc/mM). One gram of worms was Received for publication 20 May 1964. * From a dissertation submitted in partial fulfillment of the requirements for the degree Doctor of Philosophy. This investigation was supported by National Institutes of Health Training Grant No. 2E-70. t Present address: Biology Department, Rice University, Houston 1, Texas. removed after 0.5, 1.5, and 5.0 min incubation, rinsed in five changes (500 ml) of buffered Ringer's, blotted on hard filter paper, and placed in 15 ml 80% ethanol. The worms were homogenized in a blendor for 3 min at 5 C. The homogenates were centrifuged for 15 min at 1,200 g and the supernatants were extracted with 20 ml water-saturated chloroform. After centrifugation at 1,200 g for 15 min the chloroform layer was removed, the water layer was evaporated to dryness, and the residue was redissolved in 1.0 ml demineralized water. Aliquots of all extracts were plated in uniform thickness on planchets and assayed with a Baird-Atomic gas flow counter with a counting efficiency of approximately 9%. The extracts were placed on columns of Dowex 50( H+) and separated into acid effluents and NH40H eluates. These fractions were evaporated to dryness and redissolved in 0.50 ml demineralized water. Aliquots were counted as described previously. Free amino and organic acids were separated and identified by ascending oneand two-dimensional chromatography as previously described (Gr ff, loc. cit.). High-voltage electrophoresis was also employed as another means of identification of amino acids. The radioactivity in individual amino acids and organic acids was estimated by counting the radioactivity in eluates of uniform spots cut from chromatograms. Spots were located by superimposing chromatograms on fixed radioautographs over transmitted light. The 2,4-dinitrophenylhydrazine derivatives of known keto acids (pyruvate, oxaloacetate, a-ketoglutarate) and of keto acids in the acid fractions were prepared by the method of El Hawary and Thompson (1953). The hydrazones were dissolved in 0.20 ml phosphate buffer (pH 7.2). Aliquots were spotted on Whatman No. 4 paper and developed with n-butanol: ethanol: 0.5 N NH4OH

A study of the neutral lipids and phospholipids of the acanthocephala Macracanthorhynchus hirudinaceus and Moniliformis dubius

1. 1. The neutral lipids and phospholipids of male and female Macracanthorhynchus hirudinaceus and Moniliformis dubius have been studied. 2. 2. M. dubius contained some four times more lipid than M. hirudinaceus and the bulk of the higher fat content was glycerides. A sex dimorphism was observed in both species. The male worms contained almost twice the total lipid that was present in the females. 3. 3. The esterified fatty acids from the neutral lipid fractions were analyzed gas-chromatography of their methyl esters. Acids from C 10 through C 20 were present in both species and the C 18 acids accounted for over 70 per cent of the total acids. 4. 4. The phospholipids of both species contained ethanolamine, serine, choline and inositol phospholipids. Plasmalogens were also found in the phosphate containing lipids.

The Acanthor Membranes of Two Species of Acanthocephala

Acanthocephalus jacksoni and Echinorhynchus gadi acanthors have enveloping A literature review indicates many inconsistencies concerning the description and number of acanthor even within a single species. The greatest variation observed in the is in the fibrillar coat. The are more easily observed in fresh, unfixed smears of eggs. It is recommended that the traditional citing of three as the typical description be reviewed with the suggestion that are typical for the acanthor stage. Since the time of Burow's (1836) original description of three membranes, the number of reported for the acanthocephalan acanthor has ranged between and five in different species, and even within a single species. This is especially true of reports for Macracanthorhynchus hirudinaceus. Anton Meyer (1928) described the development of in this species. Yet, in a later work with this same species, von Brand (1940) refers to only three There are many similar accounts in which three are cited for this species. Most investigators have not considered the possibility of constancy of number for these Moore (1946 and 1962), in describing the number for species, Moniliformis dubius and Mediorhynchus grandis, states in the first instance that there are two distinct shells plus adhering membranes whereas, in the second instance he describes two shells and three accessory membranes. This problem has been referred to on several occasions. Manter (1928) stated that textbooks are incorrect in ascribing three eggshells to the eggs of M. hirudinaceus. Four are clearly present .... . An earlier reference was made to this problem by Korschelt and Received for publication 25 March 1964. * This study was supported, in part, by a National Defense Educational Act Fellowship and was submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Zoology at The University of New Hampshire. t Present address: Department of Biology, Suffolk University, Boston, Massachusetts. Heider (1895) in which made note of the fact that ordinarily three such embryonal are formed but that in the case of E. gigas four of them are present. Recently, Rogers (1962) has stated that they consist of three or layers. MATERIALS AND METHODS Two easily accessible species of Acanthocephala were investigated in this study to determine whether or not their mature acanthors demonstrated any basic similarities or differences in the nature of their enclosing The species, Acanthocephalus jacksoni Bullock and Echinorhynchus gadi (Zoega) Miiller, are both members of the Palaeacanthocephala. A. jacksoni was obtained from the intestine of various trout at the New Hampshire Fish Hatchery, New Hampton, New Hampshire. E. gadi was collected from the intestine of pollack and cod brought in to Gloucester, Massachusetts by commercial fishermen. The acanthors of both of these species have been described previously. Bullock (1962) described the acanthor of A. jacksoni and Hamann (1891) illustrated the acanthor of E. gadi. Both had suggested the presence of although neither cited them as such in their descriptions. Mature and gravid females of both species were collected at different seasons of the year: spring, summer, and fall for A. jacksoni and winter for E. gadi. Some were relaxed in distilled water, fixed in various fixatives, i.e., Bouin's, Helly's, and Demke's and treated with various stains. Others were sacrificed immediately upon procurement from the intestine of the definitive host. Freshly obtained females were rinsed in a Ringer's solution to remove debris usually found adhering to the body and hooks of the Acanthocephala. The worms were then blotted dry and the contents of the pseudocoel smeared onto a glass slide. The smears were allowed to air dry

Permeation of Amino Acids in Moniliformis and Macracanthorhynchus (Acanthocephala)

Both sexes of Moniliformis and Macracanthorhynchus concentrate methionine against a chemical difference. The uptake of methionine by both sexes of Moniliformis is inhibited by alanine, serine, leucine, and isoleucine-these inhibitions are reciprocal. The uptake of serine and alanine is not affected by methionine in female Macracanthorhynchus; however, methionine does inhibit the accumulation of those amino acids in males of Macracanthorhynchus. The effect of a mixture of four neutral amino acids on the uptake of methionine can be predicted by the generalized equation derived by Read et al. (1963, Ann. N. Y. Acad. Sci. 113 Art. 1: 154-205). Considerable information is available concerning the entrance of potential nutritional substances into cestodes (see the review by Read and Simmons, 1963, as well as Read, Rothman, and Simmons, 1963); however, there is a paucity of such data pertaining to that process in the Acanthocephala (Laurie, 1957, 1959). Like the cestodes, Acanthocephala have no gut and it is assumed that nutrients enter through the body wall. Read et al. presented exhaustive data concerning the initial rates of entry of a number of amino acids into Hymenolepis diminuta. In that study they demonstrated that amino acids were concentrated against a chemical gradient and that there was competitive inhibition between certain groups of amino acids. Those authors further demonstrated that there was inhibition of uptake of a given amino acid in the presence of a complex mixture of amino acids and that this competition could be predicted by an extension of the Lineweaver-Burk treatment of enzyme kinetics. The following study was undertaken: (1) to obtain information concerning the permeation of a selected group of neutral amino acids into two species of acanthocephalans and (2) to Received for publication 20 March 1964. * This research was supported in part by U. S. Public Health Service Grants AI-01384 and 2E-106. t Present address: California State College, Fullerton, California. further test the application of the generalized equation of Read et al. (vide supra). MATERIALS AND METHODS Acanthocephala used in this study were obtained from two sources. Moniliformis dubius were reared in male Sprague-Dawley rats (Holtzman Rat Co.). Cystacanths of Moniliformis were obtained from laboratory-reared and infected Periplaneta americana. Rats weighing 70 to 90 g were infected with 25 12-week-old cystacanths. The hosts were maintained in groups of ten in 24by 18by 12-inch wire cages in air-conditioned quarters: Food and water were provided ad lib. until the time of autopsy at 5 weeks. Rats were killed by a blow on the head and the small intestine removed to a container of saline composed of NaCl 120 mM, KCI 4.8 mM, CaCl2 2.6 mM, MgSO4 1.2 mM, and Tris (hydroxymethyl) aminomethane: maleate buffer 25 mM at pH 7.4. The intestine was opened with scissors and the worms gently removed from the mucosa. Macracanthorhynchus hirudinaceus were carefully dissected from swine small intestines at the Houston Packing Company and transported to the laboratory in the above saline in thermos containers at 36 to 39 C. Both species of parasites were separated according to sex and treated separately throughout the remainder of the experiments. The worms were preincubated in 30-ml beakers in the above saline for 60 min at 37.5 C. Following the preincubation period the worms were blotted and incubated in 30-ml beakers containing the amino acid solution (10.0 ml for Macracanthorhynchus, 5.0 ml for Moniliformis) for 3 min. The various C14 amino acids were used at a specific activity of 0.6 ,uc/,mole. After 3 min (-+5 sec) incubation, the worms were removed, rinsed rapidly in three changes of the saline (50 ml each), blotted, and placed in tubes containing a

Competitive Exclusion of Dactylogyrus extensus by Dactylogyrus vastator (Trematoda, Monogenea) on the Gills of Reared Carp

Carp (Cyprinus carpio Linn.) infested with D. vastator, Nybelin, 1924, are slightly or not at all infested with D. extensus (Mueller and Van Cleave, 1932). Carp infested with D. extensus were found to be free or nearly free from D. vastator. These observations were made on 796 carp in ponds and subjected to statistical analysis. Under laboratory conditions in carp infested with D. extensus and subsequently exposed to D. vastator the latter species replaces the former, i.e., the number of D. extensus on the gills falls to a minimum or to zero even when the original infection with D. extensus was heavy. Histological changes of gills infested with D. vastator result in the loss of D. extensus and, eventually, in heavy infestations of D. vastator itself. With subsequent healing, the gills again become a suitable habitat for D. extensus and D. vastator. Eventually a specific resistance to D. vastator eliminates this species from the dactylogyrid fauna of the gills, and the habitat remains suitable for D. extensus and other dactylogyrids. Antagonism between different species of parasites to which fish are naturally exposed has been discussed by Dogiel (1958) and Bauer (1959). In a review of the subject the above authors cited the following examples: an increase in infestation with Lernaea (Copepoda) is associated with diminished susceptibility to glochidia and vice versa (Wilson 1916). Infection with Eubothrium crassum (Cestoda) prevents infection with Triaenophorus nodulosus (Cestoda) in trout (Scheuring, 1923). There is also antagonism between Octobothrium melangi (Monogenea) and Clavella devastatrix (Copepoda) in Gadus merlangus (Leigh-Sharp, 1933) and between Phyllodistomum folicum (Digenea) and Myxidium lieberkiihni (Neosporidia) in the urinary bladder of Esox lucius (Petrushevski, 1940). Ivasik (1953) found that Dactylogyrus extensus and D. vastator were antagonists to Trichodina carassii (Ciliata), while the above two species of Dactylogyrus and D. anchoratus were found Received for publication 23 April 1963. * This paper is part of a doctoral thesis carried out in the Department of Parasitology, Hebrew University, Hadassah Medical School, Jerusalem, under the direction of Professor G. Witenberg. t Supported by a grant from the Fish Breeders Association, Israel. 94 together on the gills of carp and apparently showed no mutual antagonism. The above observations were made during field studies. Little experimental work has been done on interspecific competition between parasites. The few experimental demonstrations of competitive relationships include those of Adler (1954) between Leishmania infantum and Plasmodium berghei, von Brand and Tobie (1960) working with species and races of Trypanosoma, and Holmes (1961) with Moniliformis dubius (Acanthocephala), and Hymenolepis diminuta (Cestoda). MATERIAL AND METHODS My observations are based mainly on an examination of 796 carp fingerlings collected in fish ponds infested with Dactylogyrus vastator and D. extensus during the spring of 1961-1962. The total length of the fingerlings was from 20 to 50 mm. Supplementary observations were carried out on fingerlings which were transferred to the laboratory and maintained in aquaria of various sizes at two constant temperatures (21 and 28 C). The parasites were counted on gills removed quickly from the living fish and immediately pressed between two glass plates. The results were subjected to the chi-square test.

Glycogen Content in Moniliformis dubius (Acanthocephala)

Glycogen has been quantitated in Moniliformis dubius from male and female rats. It was found that male worms contained more glycogen per unit weight than female worms. This was true regardless of the sex of the host. Worms recovered from female rats were smaller than worms in the male rats. Female worms, although smaller in the female rat, contained more glycogen per unit weight than female worms from male rats. A single sex infection consisting of only male worms was accidentally obtained in a group of male rats. These worms were larger and contained more glycogen than worms from the control infection. These results suggest that there are two independent phenomena affecting worm growth and metabolism. One is the interaction of male and female worms in a single infection. The other is the effect of the endocrine system of the host on worm development and metabolism. Read (1961) reviewed the literature pertaining to carbohydrate metabolism in parasitic helminths. In recent years there has been a considerable number of investigations on this subject, especially on the cestodes. Information on carbohydrate metabolism in the Acanthocephala is still rather limited. Read and Rothman (1958) investigated the carbohydrate requirements of Moniliformis dubius. They found that normal growth of this worm was dependent on the presence of carbohydrate in the host diet. They further reported that the glycogen levels in M. dubius showed a diurnal fluctuation which was correlated with the feeding time of the host. The glycogen content in M. dubius was also affected by the type of carbohydrate in the host diet. We have carried out additional quantitative studies of the glycogen content in M. dubius. The results show that other variables besides diet and feeding time of the host influence the quantity of glycogen in this worm. MATERIALS AND METHODS Male and female Sprague-Dawley rats, 3 months old at time of infection, were used as hosts. Cystacanths of M. dubius were obtained from laboratory-infected roaches. Each rat was infected with 20 cystacanths by the method of Moore (1946). Unless stated otherwise, the worms used in this study were collected 39 days after infection. Infected rats were starved 24 hr prior to autopsy. This starvation time was selected in an attempt to insure a standardized level of endogenous carbohydrate in Moniliformis. The rats were killed by a sharp blow on the head. The worms were then washed in Ringer's solution, blotted on filter paper, and weighed on a torsion balance. They were then used for glycogen analysis. The rats used in this study were housed in cages 18 by 24 by 10 inches. Each cage contained 12 animals. A vertical feeding tray and two water bottles were provided for each cage. The rats were allowed to feed ad libitum. Under these conditions the recovery of adult worms from a 20cystacanth infection averaged about 70% (49% to 100%). This recovery agreed with that reported by Burlingame and Chandler (1941) for M. dubius during a 39-day infection period. Glycogen was determined by the method of Carroll et al. (1956). The worms were homogenized in 5% trichloroacetic acid, and the homogenate was centrifuged. Glycogen was precipitated from the supernatant by the addition of ethanol to a final concentration of 70%. The resulting precipitate was collected by centrifugation, washed in 70% ethanol, and recentrifuged. The precipitate was then dissolved in water to a desired volume. Aliquots were analyzed for carbohydrate by use of the anthrone reagent of Roe (1955). The color was measured on a Coleman Junior Spectrophotometer at 620 mjz. Appropriate mean values for worm weight and glycogen content were compared by Student's t test. The t values are given in the text. Differences between means were considered significant when P < 0.05. Received for publication 24 October 1962. * This work was supported by National Institutes of Health Training Grant No. 2E-70 and University of California Faculty Research Grant No. 1718 (Zoology). 204 This content downloaded from 157.55.39.249 on Wed, 03 Aug 2016 06:01:54 UTC All use subject to http://about.jstor.org/terms GRAFF AND ALLEN-GLYCOGEN IN MONILIFORMIS 205 TABLE I. Wet weight and glycogen content of Moniliformis dubius from male rats.

Establishment and distribution of Raillietina cesticillus in the fowl and comparative studies on amino acid metabolism of R. cesticillus and Hymenolepis diminuta

1. 1. The establishment and distribution of Raillietina cesticillus in White Rock chickens was followed for 0, 1, 2, 5, 8, 11, 14, and 17 days after inoculation. The cestode was found first to establish anterior to the Ampulla of Vater. By the third day the tapeworm was distributed along the first 14 centimeters posterior to the opening of the bile ducts, where it remained during its adult life in the intestine. These movements were compared to those of Hymenolepis diminuta and Moniliformis dubius. 2. 2. The free amino acid and protein fractions of Raillietina were analyzed by paper chromatography, and compared to analyses of Hymenolepis. The latter worm contained a greater amount of free amino acid than the former and an especially larger amount of taurine. Raillietina had a particularly large amount of proline. The protein hydrolysates of the two worms were very similar. 3. 3. The following transaminase reactions were found in Raillietina, listed in order of decreasing activity: glutamate to aspartate and aspartate to glutamate; alanine to glutamate and glutamate to alanine (with oxalacetic acid); and glutamate to alanine. In comparison to the activity of H. diminuta, determined directly and by previous studies, the activity of the two most active reactions in Raillietina was approximately one-half that found in Hymenolepis. In addition, Hymenolepis also possessed an equally active alanine to glutamic acid system as well as the two systems mentioned above. 4. 4. The importance of the particular position of worms in the intestine was discussed, and the significance of the transaminase reactions to cestodes was pointed out.

Aerobic metabolism of Moniliformis dubius (Acanthocephala)

Moniliformis dubius synthesized glycogen from glucose, fructose, mannose, and maltose. The synthesis of trehalose from these substrates was not demonstrated although this disaccharide was present in considerable amounts. Glycogen and trehalose were the major endogenous carbohydrates. Acids excreted during the aerobic fermentation were identified tentatively as acetic, lactic and formic acids. These were formed in the presence or absence of exogenous carbohydrates. Glucose, fructose, mannose, and maltose increased the rate of lactic acid excretion. Mannose, maltose and possibly fructose stimulated respiration. Glucose and glycerophosphoric acid did not.

Histochemical studies on the Acanthocephala. III. Comparative histochemistry of alkaline glycerophosphatase

1. 1. The distribution of alkaline glycerophosphatase has been studied in 23 species, 18 genera, 8 families, and all three orders of the phylum Acanthocephala. A standardized form of the Gomori technic was used in this study. 2. 2. When present, phosphatase activity was most pronounced in the outer layer of the subcuticula of the trunk. Considerable enzyme was also present in the inner layer of this subcuticula. The middle layer of the subcuticula of the trunk and most other organs were negative. The proboscis and lemnisci were usually negative but a marked reaction was observed in Fessisentis vancleavei. 3. 3. Enzyme activity was observed in the following adult worms: Illiosentis furcatus, Telosentis tenuicornis, Polymorphus sp., Corynosoma wegeneri, Fessisentis vancleavei, Echinorhynchus gadi, Pomphorhynchus bulbocolli, Moniliformis dubius, and Macracanthorhynchus hirudinaceus. Larvae and juveniles of C. wegeneri, Arhythmorhynchus duocinctus, Fessisentis vancleavei, and Oncicola canis were negative except for a slight lemniscal reaction in the latter. Cystacanths of M. dubius showed phosphatase in the inner layer of the subcuticula while an immature Macracanthorhynchus ingens had some activity in the lemnisci and the subcuticula. 4. 4. Eight members of five genera of the Neoechinorhynchidae were alwavs negative for alkaline phosphatase. Leptorhynchoides thecatus and Acanthocephalus sp. were also devoid of enzyme that could be demonstrated by the technic used. 5. 5. The enzyme in the outer layer of the subcuticula is probably associated with absorption or secretion from the body surface. The function of the phosphatase of the inner layer is not clear. 6. 6. Those forms which were phosphatase negative in long series of worms under different physiological conditions probably use other enzyme systems in their absorptive or secretive processes.

The in vitro fermentation of carbohydrates by two species of cestodes and one species of acanthocephala

Under anaerobic conditions, the tapeworms, Hymenolepis diminuta and Oochoristica symmetrica, ferment glucose and galactose, at least partially, to acidic end-products. The fermentation of mannose and xylose by the former and of mannose by the latter is somewhat equivocal. The acanthocephalan, Moniliformis dubius, ferments glucose, galactose, mannose, fructose and maltose to acidic catabolites. No evidence of the acidic fermentation of a variety of other carbohydrate substrates by these helminths has been found. Phlorizin, non-competitively, inhibits the fermentation of glucose by H. diminuta but has no effect on the fermentation of glucose, mannose, fructose, and maltose by M. dubius. Dibromophlorizin inhibits glucose and galactose fermentation by H. diminuta but has no influence on sugar fermentation by M. dubius. These glucosides inhibit galactose fermentation but not glucose fermentation by O. symmetrica. The fermentation of glucose and fructose by M. dubius is inhibited by p-chloromecuribenzoic acid but not by iodoacetic acid. Quinacrine inhibits the fermentation of glucose and galactose by O. symmetrica and glucose fermentation by H. diminuta; whereas, acriflavine inhibits glucose fermentation but not galactose fermentation by these cestodes.

Effect of gonadectomy and gonadal hormones on singly established Hymenolepis diminuta in rats

The effects of gonadectomy and gonadal hormones on singly-established H. diminuta in rats on “complete” and “deficient” diets have been studied by means of estimations of the number of eggs eliminated by the parasite per 24-hour periods over varying intervals of time. A decline in the number of eggs of H. diminuta eliminated per 24 hours over a period of 3 months was observed in castrated male rats fed a “complete” diet which was comparable to the effects observed in normal male hosts on a “deficient” diet. Castrated males fed a “deficient” diet, however, exhibited a decline in the numbers of eggs of this cestode eliminated which was comparable to that observed in normal female hosts on a “deficient” diet. Testosterone and progesterone, administered at the rate of 1 mg per day, restored egg output to approximately normal levels in castrated male hosts. Testosterone was also similarly effective in normal female rats on “deficient” diet, but progesterone was ineffective. Chorionic gonadotropin raised the egg output of H. diminuta in normal male and female rats on “deficient” diet but not in gonadectomized hosts. A stimulation of testosterone production is postulated as a possible mechanism in the case of normal male rats and possibly the production of estrogen in the normal female hosts. Accidental concurrent infections with a few Moniliformis dubius had no evident effect on egg production of H. diminuta. These acanthocephalans were lost quickly from hosts on “deficient” diet.

Studies on the Life History and Development of Moniliformis dubius Meyer, 1933

Grassi and Calandruccio (1888) reported the occurrence of Echinorhynchus moniliformis (Moniliformis moniliformis) in the small intestine of Mus decumanus and Myoxus quercinus in Sicily. They found the common beetle Blaps mucronata to be the intermediate host, and three times the authors found more than one hundred juvenile E. moniliformis in the body cavity of a single Blaps. They fed several of these juvenile acanthocephalans to a young rat and two weeks later recovered worms 1 cm long. Calandruccio proved the susceptibility of man to infection with this acanthocephalan by ingesting the juveniles recovered from the beetle. Five weeks were required for the worms to reach sexual maturity. Magalhaes (1898) reported the occurrence of the juveniles of Moniliformis moniliformis (M. dubius) in the body cavity of Periplaneta americana collected in Rio de Janeiro. His identification of these larvae was made on the basis of shape, size, and distribution of the proboscis hooks, and was not verified by experimental feeding. Travassos (1917) found the juveniles of Moniliformis dubius (M. travassosi Meyer, 1933) in the body cavity of Periplaneta americana in Brazil. In preliminary attempts to infect cockroaches with Moniliformis dubius, Chandler (1921) reported negative results. However, in later work, investigators in Chandler's laboratory (Chandler, 1941) have had no difficulty in infecting P. americana with M. dubius.