Volume 56, Issue 4

Random amplification of polymorphic DNA polymerase chain reaction (RAPD-PCR) was carried out to assess the genetic diversity of five new microsporidian isolates viz., NIWB-11bp, NIWB-12n, NIWB-13md, NIWB-14b and NIWB-15mb identified from the silkworms. A type species, NIK-1s_mys was used as control for comparison. Differences in the spore shape, length and width were observed. Of the 30 decamer random primers tested, 22 primers gave repeatable RAPD profiles and yielded a total of 143 fragments, of which 78 were polymorphic (55%). The resulting data was used to derive genetic similarity values for constructing a dendrogram. The neighbour joining method based on Dice coefficients indicate a major cluster comprising NIK-1s_mys, NIWB-11bp and NIWB-12n, whereas NIWB-13md, NIWB-14b and NIWB-15mb appear to be different from each other as well from the major cluster mentioned above which includes the type species (NIK-1s_mys). Based on the reproducibility of RAPD profiles, we are able to identify these microsporidians as different isolates. The RAPD technique may be useful in detecting sources of infection of this economically important domestic insect.

An undescribed species of Microsporidia Balbiani, 1882 was isolated from the muscularis mucosa of the intestinal mucosa of the reared Atlantic bluefin tuna (Thunnus thynnus). Transmission electron microscopy showed that the pyriform unikaryotic microsporidium, measuring 3.6 μm ± 0.08 in length and 1.8 μm ± 0.04 in width (inferred from TEM sections), had two layers of 14–17 coils of polar filament and a robust manubrium of the filament. No developmental stages were observed in xenoma. Phylogenetic analysis of the small subunit rRNA showed closest similarity with Kabatana spp. (88.4%), clustering the microsporidium together in a sister group with K. takedai and Kabatana sp. (JI-2008)/K. newberryi clade. While recognized Kabatana members show ovoid, rounded to pyriform spore, lack of sporophorous vesicle or xenoma, 3–10 coils of polar filament in 1–2 rows and are localized within cytoplasm of trunk muscle, the new species is markedly pyriform, xenoma forming, with many filament coils in 2 rows and parasitizing smooth intestinal muscle. Since morphological features were not typically congruent with any of Kabatana spp. so far described and molecular clustering indicated paraphyletic position within Kabatana clade, we suggest assigning described species to collective group Microsporidia, as Microsporidium milevae sp. nov., until more evidence permits potential formation of the new genus.

Microsporidia are intracellular opportunistic parasites that cause chronic diarrhea in AIDS patients. Little is known about the prevalence of these pathogens in pediatric cases with cancer and diarrhea. Unidentified causes of chronic diarrhea were previously encountered in pediatric cancer patients at the National Cancer Institute in Egypt. Therefore, this study tried to search for the contribution of microsporidia as a causative agent of diarrhea in this population using acid-fast trichrome stain as a specific staining and the PCR in order to evaluate the staining technique in clinical diagnosis of microsporidia. Between January 2008 and June 2009, 271 diarrheic samples from pediatric patients with cancer were studied. Microsporidia were confirmed in 13 (4.8%) cases by both PCR and staining, and additional 2 samples were positive only by staining. As a negative control, stool samples from 60 diarrheic children without malignant cancer and no microsporidia detection were examined by these two methods. So, it can be concluded that adding a diagnostic test for microsporidia to the clinical laboratory work in hospitals concerned with cancer is essential. Acid fast trichrome staining technique as being nearly efficacious as the PCR, but simpler and less expensive, can replace the molecular techniques for the diagnosis of microsporidia.

Schizophrenia is a serious neuropsychiatric disease of uncertain etiology. Previous studies have demonstrated that antibodies to Toxoplasma gondii infection are associated with an increased risk of schizophrenia. The objective of this study was to analyze anti-T. gondii antibodies in 477 Chinese schizophrenia patients using an enzyme-linked immunosorbent assay (ELISA) based on recombinant surface antigen 1 (rSAG1), recombinant bradyzoite antigen 1 (rBAG1) and the soluble tachyzoite antigens (STAg) of T. gondii RH strain. Results showed that among the sero-positives (IgG and/or IgM) for T. gondii infection examined in schizophrenia patients, sero-positive samples for rSAG1, rBAG1 and STAg were 20.5% (98/477), 20.5% (98/477) and 23.5% (112/477) respectively, while compared to 210 blood donors, sero-positive (IgG and/or IgM) samples for these antigens (rSAG1, rBAG1 and STAg) were only 5.7% (12/210), 6.2% (13/210) and 5.7% (12/210), respectively. Furthermore, when IgG antibody reaction in the schizophrenia sera was compared with the rBAG1 and rSAG1, results demonstrated that beside the cases which can be detected by both rSAG1 and rBAG1, some sero-positive for T. gondii in schizophrenia sera can only be detected either by rSAG1 or rBAG1. This phenomenon was also observed in the detection of IgM with rSAG1 and rBAG1. 5.9% (28/477) of cases of schizophrenia which are positive for IgG or IgM by rSAG1 are negative for STAg, while 9.2% (44/477) of the schizophrenia cases which are positive for IgG or IgM by rBAG1 are negative for STAg. Although STAg can also be used to diagnose T. gondii infection from schizophrenia patients, it may not actually indicate the infection as some positive samples may be mistakenly considered to be negative. In conclusion, our results demonstrate that the sero-positive rate for T. gondii in the Chinese schizophrenia patients was higher than blood donors. More importantly, our results provide evidence that the combination of rSAG1 and rBAG1 antigens in the diagnosis of T. gondii infection could closely reflect the actual infection of this parasite in schizophrenia patients.

A new genus and species of Gyrodactylidae, Ieredactylus rivuli gen. et sp. nov. (Platyhelminthes, Monogenea), is described from the skin of Hart’s Rivulus (Rivulus hartii Boulenger), a cyprinodontiform fish collected from streams of the Caroni and Oropouche drainages and the Pitch Lake in Trinidad (prevalence all localities: 16.7–94.6%; mean parasite intensity 1–9 parasites/fish; range 1–34) with the holotype originating from a tributary of the Aripo River. This viviparous monogenean is distinctive from other genera of Gyrodactylidae by its split ventral bar membrane, the shape of its male copulatory organ, the presence of two conical accessory pieces associated with the hamulus root and two differently shaped marginal hook sickles. Its unique rDNA sequence shows the closest ITS2 similarity (70%) to Gyrodactyloides andriaschewii Bychowsky et Poljansky, 1953. The presence of I. rivuli gen. et sp. nov. in the Pitch Lake indicates an adaptation to extreme environmental conditions such as high temperatures and hydrocarbons and adverse pH. Guppies may potentially serve as temporary hosts. The parasite displays distinct behaviours, including a characteristic ‘swimming-like’ movement. The ecology and phylogeny of I. rivuli gen. et sp. nov. is discussed in relation to the diversity of other gyrodactylids in Trinidad.

Spermatophores are described in the microbothriid monogenean Dermopristis cairae from the giant shovel-nosed ray, Glaucostegus typus (Elasmobranchii, Rhinobatidae). Each spermatophore consists of a fusiform capsule containing sperm and a hollow stalk. The proximal ends of the stalks of fully formed spermatophores were open and located in a recess on the ventral surface close to, but not inside, the vagina and the male reproductive opening. Three adult parasites carried spermatophores attached externally to the ventral surface. One individual carried 3 spermatophores and 2 others a single spermatophore. In addition, in 2 adults, short lengths of what appeared to be stalk remnants were observed attached near the reproductive openings. With regard to spermatophore exchange, the evidence in support of the following 2 possibilities is discussed. (1) Adult specimens of D. cairae carrying fully formed spermatophores ventrally are recipients not donors, having received their spermatophores singly during mating events. (2) Spermatophore carrying adults are donors, their spermatophores being freshly made and on offer to a potential mate. The evidence points to the first of these alternatives as the most likely but this is not conclusive and requires observations on mating between living parasites.

Maritrema portucalensis sp. nov. (Digenea, Microphallidae) from Aveiro estuary, northern Portugal, is described on the basis of excysted metacercariae obtained from the gill lamellae of the shore crab Carcinus maenas (L.). M. portucalensis sp. nov. most closely resembles Maritrema subdolum Jägerskiöld, 1909, a species that also frequently uses C. maenas as second intermediate host. The new species differs from M. subdolum in having a narrower body at the level of testes and a smaller ovary and testes. Also, the digestive caeca presented various folds along their length, thickening in the terminal region. Moreover, the body surface was fully covered by tegumental spines provided with several teeth. To complement our morphological characterization and to identify M. portucalensis metacercariae, the ITS1 rDNA region of the cysts isolated from C. maenas was sequenced and compared with the corresponding available sequences of digenean trematodes belonging to the family Microphallidae. Alignments revealed 28 base-pair differences between the query-ITS1 and that of M. subdolum, and more importantly a 100% similarity with the sequence of Microphallidae sp. no. 15 cercaria from the snail Hydrobia ulvae. The last observation allowed us to establish an unequivocal association between cercaria known as Microphallidae sp. no. 15, and the metacercaria of M. portucalensis sp. nov. characterized in this work, contributing decisively to the clarification of its life cycle.

Two cercariae, one of them ocellate and with well developed tail (Monorchiidae) and another apharyngeate brevifurcocercous (Aporocotylidae), parasite of Amiantis purpurata (Lamarck, 1818) (Bivalvia, Veneridae) from the Patagonian coast on the Southwestern Atlantic Ocean, are described. These reports comprise the second monorchiid intramolluscan infection reported for the Southern Hemisphere and first intramolluscan aporocotylid for the Southern Hemisphere. In addition, this constitutes the first report of aporocotylid intramolluscan stages (parthenita) occupying only the haemocoel of the gills of a marine molluscan host rather than the digestive gland and gonad, the usual site of infection.

Vitellogenesis and vitellocytes of the bothriocephalidean cestode Clestobothrium crassiceps (Rudolphi, 1819), a parasite of the teleost fish Merluccius merluccius (L., 1758), were studied by means of transmission electron microscopy (TEM) and cytochemistry. During vitellogenesis, four developmental stages were distinguished at the TEM level: (I) a stem cell stage of the gonial type; (II) an early differentiation stage, predominantly exhibiting lipid and protein synthetic activity; (III) an advanced differentiation or vitellocyte maturation stage, primarily exhibiting active glycogenesis still accompanied by an increase in lipid accumulation; and (IV) a mature vitellocyte stage. Vitellogenesis involves: (1) an increase in cell volume; (2) an extensive development of parallel, frequently concentrically arranged, cisternae of granular endoplasmic reticulum (GER) that produce dense, proteinaceous shell-gobules; (3) the development of Golgi complexes engaged in the packaging of this material; (4) an accelerated accumulation of unsaturated and saturated lipid droplets, along with their continuous enlargement and fusion; (5) the formation of individual β-glycogen particles and α-glycogen rosettes and their accumulation in the form of glycogen islands scattered among lipid droplets in the cytoplasm of maturing and mature vitellocytes; and (6) the rapid accumulation of large, saturated lipid droplets accompanied by dense accumulations of α- and β-glycogen along with proteinaceous shell-globules or shell-globule clusters in the peripheral layer during the advanced stage of vitellocyte maturation. Vitellogenesis in C. crassiceps generally resembles that previously described for three other bothriocephalideans, but differs from that of other cestode orders. Cytochemical staining with periodic acid-thiocarbazide-silver proteinate for glycogen indicates a strongly positive reaction for β-glycogen particles and α-glycogen rosettes, which form several large glycogen accumulations around the large, saturated lipid droplets of maturing and mature vitellocytes. Some hypotheses concerning the interrelationships between patterns of vitellogenesis, the possible modes of egg formation, embryonic development and life cycles in cestodes, and their phylogenetic implications are commented upon.

Rhabdias mcguirei sp. nov., is described on the basis of specimens found in the lungs of northern Philippine flying lizards, Draco spilopterus (Reptilia, Agamidae) collected in Aurora province, Luzon Island, Philippines. It is characterized by a rounded oral opening, a buccal capsule consisting of anterior and posterior parts, and the shape of the cuticular inflation in the anterior part of the body: the cuticle is less inflated in the anterior-most part, with the inflation gradually thickening up to the level of the oesophageal-intestinal junction. The new species is differentiated from the 11 most closely related species of Rhabdias previously known from lizards.

The large stomach worm, Haemonchus contortus is a highly pathogenic nematode parasite of sheep (Ovis aries) and goats (Capra hircus). It sucks blood and causes acute disease, anaemia and severe economic losses due to morbidity and mortality. This paper compares characteristics of surface cuticular ridges (synlophe) and morphometrics of total body length, cervical papillae, spicules, barbs, gubernaculum and vulvar flap morphology of H. contortus recovered in the two hosts from the States of Perak and Kelantan, Peninsular Malaysia. The morphological variation in H. contortus samples recovered from the two hosts in Kelantan was found to be higher than from Perak. A discriminant function, based on three measurements of the spicule, confirmed the identity of male worms in the two hosts as belonging to the same species H. contortus.

Five species of nematode were collected from the digestive tracts of 10 Coccymys ruemmleri from Papua New Guinea. Montistrongylus giluwensis sp. nov. can be distinguished from its congener by its size, in having more ridges in the synlophe, longer spicules and the form of the cuticle on the female posterior end. Syphacia (S.) coccymyos can be distinguished from all other species in the subgenus by having a laterally extended dorso-ventrally constricted oval cephalic plate and both cervical and lateral alae, together with the combination spicule length and egg size. The helminth assemblage had similar species richness to that of Chiruromys vates with Odilia mackerrasae the only species in common.

Lemuricola (Madoxyuris) vauceli Chabaud, Brygoo et Petter, 1965 is redescribed on the basis of a re-examination of type and voucher specimens. Using both light and SEM microscopy, structures such as the cephalic papillae, details of the lateral alae, area rugosa and tail cuticular ornamentation, are described or redescribed with more precision. The detailed measurements of several male and female specimens are given in a table. In the original description, using light microscopy only, the authors had described 3 pairs of pedunculated papillae in the male of L. vauceli. The present study establishes the precise number, shape and disposition of the end of the male tail to include only two pairs of pedunculated papillae, 2 pairs of sessile papillae, and 1 pair of phasmids. As the same pattern was observed in all descriptions and recent redescriptions of species classified in the Enterobiinae, the present work confirms that the number and disposition of the caudal papillae observed in the subfamily is constant.

Parasitological examination of the freshwater fish Tetraodon palembangensis Bleeker (Tetraodontidae, Tetraodontiformes) (n = 19), imported along with other ornamental fishes from Thailand into the Czech Republic in October 1998, revealed the presence of male specimens of the nematode Dichelyne (Dichelyne) hartwichi Moravec, Wolter et Körting, 1999 (Cucullanidae) (prevalence 11%, intensity 1). Their detailed study using light and scanning electron microscopy (the latter used for the first time in this species) enabled the redescription of this hitherto little-known species and the recognition of considerable morphological and biometrical intraspecific variability. The finding of this species in T. palembangensis represents a new host record.

Over 700 specimens of Corynosoma strumosum (Rudolphi, 1802) Lühe, 1904 were collected from one young male Caspian seal, Pusa caspica (Gmelin) in the southern land-locked Caspian Sea in April, 2009. Collected worms showed consistent variations from those reported by other observers using light microscopy especially in proboscis hook and trunk spine patterns. SEM images revealed many features that have not been previously reported including the shape and distribution of trunk spines, dorsoventral differences in proboscis hooks and their organization, the baldness of anterior proboscis, the rough egg topography, epidermal micropores, and variations in the female gonopore. This isolated population of C. strumosum from the land-locked Caspian Sea is distinguished from others reported from open waters elsewhere by the distribution of trunk spines, consistently smaller size of trunk and testes, larger eggs, and fewer proboscis hooks. Histopathological sections reveal the invasive path of worms in host tissue with damage to intestinal villi and worm encapsulation. Information obtained from SEM studies and histopathological sections is reported for the first time.

A checklist of the parasitic Cymothoidae of Indian fishes was compiled from parasitological records published between 1783 and 2011. The checklist is arranged alphabetically, providing valid names, synonyms and authorities of the parasite species, as well as valid names and synonyms of the host fish, its capture sites, author(s) and date of published records. The host list consists of all parasites species listed under the host species. A total of 47 nominal species corresponding to 36 valid species are listed from 74 host species belonging to 34 families. Several parasites not identified to species level and parasite species without the host data or where the parasite was found not associated with a fish are also included in this checklist.