The two flies Fannia canicularis Linn, and F. scalaris Fab. are, on account of their habits, of considerable economic importance in their relation to man. It is therefore desirable that those engaged in public health and medical work and others should have a knowledge of the breeding and other habits of these flies, which they are certain to meet in their work under circumstances of varying importance. The inquiry of which this account is the result was undertaken several years ago at the request of Dr Monckton Copeman in connection with the Local Government Board's inquiry on the carriage of infection by flies in the reports of which a portion of this paper has been included. Owing to my removal from England to Canada in 1909 and subsequent pressure of work, its completion was delayed.

Aphanomyces astaci causes crayfish plague, which is a devastating disease of European freshwater crayfish. The likely first introduction of A. astaci into Europe was in the mid-19th century in Italy, presumably with the introduction of North American crayfish. These crayfish can carry A. astaci in their cuticle as a benign infection. Aphanomyces astaci rapidly spread across Europe causing the decline of the highly susceptible indigenous crayfish species. Random amplified polymorphic DNA-PCR analysis of A. astaci pure cultures characterized five genotype groups (A, B, C, D and E). Current A. astaci genotyping techniques (microsatellites and genotype-specific regions, both targeting nuclear DNA) can be applied directly to DNA extracted from infected cuticles but require high infection levels. Therefore, they are not suitable for genotyping benign infections in North American crayfish (carriers). In the present study, we combine bioinformatics and molecular biology techniques to develop A. astaci genotyping molecular markers that target the mitochondrial DNA, increasing the sensitivity of the genotyping tools. The assays were validated on DNA extracts of A. astaci pure cultures, crayfish tissue extractions from crayfish plague outbreaks and tissue extractions from North American carriers. We demonstrate the presence of A. astaci genotype groups A and B in UK waters.

The sheep scab mite, Psoroptes ovis, induces an intensely pruritic exudative dermatitis which is responsible for restlessness, loss of appetite and weight loss. Within the first 24 h of infection, there is a rapid inflammatory influx of eosinophils and apoptosis of the keratinocytes at the site of infection. The former cell type is capable of a sustained respiratory burst, toxic products of which may directly damage the mite and also contribute to lesion formation. Analysis of a P. ovis expressed sequence tag (EST) database identified a number of antioxidant enzyme-encoding sequences, including peroxiredoxin (thioredoxin peroxidase EC 1.11.1.15), all of which may help the mite endure the potentially toxic skin environment. A full length sequence encoding Po-TPx, a protein of 206 amino acids which showed high homology to a peroxiredoxin from the salivary gland of the tick Ixodes scapularis, was amplified from P. ovis cDNA. Recombinant Po-TPx was expressed in bacteria and antiserum to this protein was used to localize native Po-TPx in mite sections. Peroxiredoxin was localized, amongst other sites, to a subpharyngeal region in mite sections. The recombinant protein was recognized by sera from sheep infested with the mite suggesting that it may be secreted or excreted by the mite and interact with the host immune response.

Ocular toxoplasmosis (OT) caused by Toxoplasma gondii is a major cause of infectious uveitis, however little is known about its immunopathological mechanism. Susceptible C57BL/6 (B6) and resistant BALB/c mice were intravitreally infected with 500 tachyzoites of the RH strain of T. gondii. B6 mice showed more severe ocular pathology and higher parasite loads in the eyes. The levels of galectin (Gal)-9 and its receptors (Tim-3 and CD137), interferon (IFN)-γ, IL-6 and IL-10 were significantly higher in the eyes of B6 mice than those of BALB/c mice; however, the levels of IFN-α and -β were significantly decreased in the eyes and CLNs of B6 mice but significantly increased in BALB/c mice after infection. After blockage of galectin–receptor interactions by α-lactose, neither ocular immunopathology nor parasite loads were different from those of infected BALB/c mice without α-lactose treatment. Although the expressions of Gal-9/receptor were significantly increased in B6 mice and Gal-1 and -3 were upregulated in both strains of mice upon ocular T. gondii infection, blockage of galectins did not change the ocular pathogenesis of genetic resistant BALB/c mice. However, IFN-α and -β were differently expressed in B6 and BALB/c mice, suggesting that type I IFNs may play a protective role in experimental OT.

Post-kala-azar dermal leishmaniasis (PKDL) is an unusual dermatosis that develops as a sequel in 5–15% of cured cases of kala-azar (KA) after months or years of treatment in India. Molecular differences are reported to exist between the KA and PKDL isolates which may underlie the diversity in clinical manifestations of the disease. Here, arbitrary primed-PCR (AP-PCR) has been used for genetic fingerprinting of parasite isolates from dermal lesions of PKDL patients (n=14) and compared with bone-marrow derived parasites from KA patients (n=3). All isolates showed an identical AP-PCR pattern with 4 arbitrary primers. Further, AP-PCR was exploited to identify the stage regulated genes of the parasite. Six polymorphic fragments were identified in PKDL in comparison with KA isolates, and were subjected to Northern blot analysis. Five polymorphic fragments represented transcribed sequences; 4 out of 5 drew differential expression in pro- and amastigote stages, although the expression was comparable between PKDL and KA isolates. The study led to the identification of genes, which exhibit stage-regulated expression in Leishmania donovani derived from PKDL or KA patients, including a putative phosphodiesterase, DEAD box RNA helicase, iron superoxide dismutase b (fesodb) and a hypothetical protein. Demonstration of transcripts of DEAD box RNA helicase in PKDL and KA diseased tissues implicates its role in disease pathogenesis.

The dry weight, nitrogen content, and total carbohydrate content of Hymenolepis diminuta were studied after a period of 15 days growth in rats, maintained on different diets. The special diets were fed during the last 7 days of this period.

Using equal iso-molecular quantities, seven amino acids, representing the different classes of neutral amino acids, acidic amino acids and basic amino acids, have been screened for their effect on worm growth when added as free dietary supplement to the different diets. All of them, to a greater or lesser extent, had significant effects on worm growth.

The award of a research grant by the Standing Advisory Committee for Medical Research in the British Caribbean is gratefully acknowledged. I am also indebted to Professors J. C. Waterlow and C. von Holt for the loan of equipment in order to carry out the carbohydrate and nitrogen determinations, to Professor I. M. Good-body for his continued interest and support, and to the Rippel Foundation for research facilities.

1. The morphology and life history of a gregarine Lankesteria culicis (Ross), as it lives in the gut of Aëdes (Finlaya) geniculatus (Olivier), are described. The earlier stages are intra-epithelial in the fore and anterior mid-gut of the larvae. After a period of growth the trophozoites are liberated into the gut lumen where they attach themselves to the epithelium by a well-developed epimerite which functions as a sucker. Cyst formation and further stages in sporogony take place in the malpighian tubules when the larvae develop into the pupae. In the adult mosquito only ripe sporocysts are found, packed in the malpighian tubules and scattered in the hind-gut. The spores escape through the alimentary canal.

2. The systematic position of Lankesteria is reviewed. In view of its true dicystid nature, combining both acephaline and cephaline features, it is suggested that its proper systematic position would be in the dicystid family Lecudinidae Kamm emend. Reichenow, 1929, which includes all forms intermediate in position between the monocystids and the true tricystids.

Trichinous mice were killed and their carcasses maintained at room temperature or subzero temperatures for varying lengths of time. Some worm parameters were measured after direct isolation from carcasses while others were measured following passage through a second round of hosts. Glycogen and trehalose levels in infective 1st-stage larvae (L1) isolated directly from carcasses maintained at room temperature were significantly less than controls (day 0) after day 4 post-kill (p.k.). By day 21 p.k. among L1 isolated directly from mouse carcasses those observed coiled or moving had decreased by around 20% compared to day 0 p.k. The percentage of L1 isolated from carcasses on several days p.k., used to infect mice and recovered as pre-adult worms declined significantly after they had remained in carcasses for 14 days. Only 2·6% of muscle larvae isolated from carcasses on day 21 p.k. and used to infect mice were recoverable as pre-adults. Pre-adult worms raised in mice infected with larvae from day 7 carcasses had about 50% less glycogen than worms raised in mice infected with larvae isolated from fresh carcasses (day 0 p.k.). The fecundity in vitro of adult worms isolated from mice on day 7 following infection with infective L1 larvae maintained in carcasses held at room temperature for 0–16 days declined only when adult worms developed from larvae recovered from carcasses at 3 days following host death. Following 24 h at temperatures below freezing, infectivity of L1 larvae isolated from frozen carcasses was reduced by 97%. Carbohydrate levels remained high in larvae from carcasses frozen for up to 4 days.

Parasites with complex life cycles face 2 major challenges for transmission in northern latitudes. They have to cope with the general unpredictability associated with the series of transmission events required for completion of the cycle, and transmission has to be completed within a narrow temporal window because of strong seasonality. Despite this, some parasites show high transmission success, suggesting the operation of effective transmission mechanisms. We explored the transmission of Ichthyocotylurus pileatus (Trematoda) from its snail (Valvata macrostoma) to fish (Perca fluviatilis) hosts by examining some key characteristics in the dynamics of the cercarial emergence from snails. Transmission took place within a few weeks mainly in July, thus verifying the narrow temporal window for transmission. The output of the short-lived cercariae from the snails was low and variable in magnitude, but nevertheless resulted in a rapid and high rate of infection in newly hatched fish. The cercarial emergence showed a strong circadian rhythm with most of the cercariae emerging in early evening and night, which might represent the most likely mechanism underlying the high rate of transmission in this species. We emphasize the importance of holistic approaches combining aspects of multiple host species in studies on transmission of complex life-cycle parasites.

Malate dehydrogenase (decarboxylating) from Tritrichomonas foetus hydrogenosomes was purified close to homogeneity by a combination of differential centrifugation, zwitterionic detergent solubilization, Red-Sepharose chromatography and anion-exchange chromatography. The enzyme with apparent subunit size of 59 kDa and native molecular mass of 308 kDa utilized NAD+ preferentially to NADP+ as a cofactor and required Mn2+ or Mg2+ for its activity. Affinity curves for malate and coenzymes were hyperbolic. Km for malate was 100 μM and 458 μM in the presence of NAD+ and NADP+, respectively. Km for NAD+ and for NADP+ in the presence of malate was 18 μM and 207 μM, respectively. The enzyme is proposed to be a tetramer with a possible physiological role in the maintenance of an appropriate NAD+/NADH ratio in hydrogenosomes.

The epidermis of Diplectanum aequans has, in general, been found to be similar to the epidermis of other monogeneans, consisting of a syncytial outer epidermis and sunken sub-epidermal nucleated regions. However, the epidermis of D. aequans differs from that of other monogeneans in 3 respects. These are, the presence of large areas of granular cytoplasm within the outer epidermis, the presence of myofibres invaginating into the epidermal matrix and, in the posterior regions of the epidermis, the presence of epidermal scales. These scales occur within the epidermal cytoplasm, beneath the outer membrane, and are composed of moderately electron-dense material. Also present beneath the outer membrane in the more anterior regions of the epidermis are small scale-like sclerites of a similar electron density to the epidermal scales.

The ability of certain species of trypanosomes (T. lewisi and T. equiperdum) to ingest previously stained oil drops which occurred simultaneously in the blood has been observed.

Tryparsamide incorporated in dyed (Sudan IV) cod-liver oil.when administered orally, was found to be effective against T. equiperdum infections in mice.

Here we describe extracellular matrix alterations in footpad lesions and draining lymph nodes caused by Leishmania (L.) amazonensis in mouse strains with distinct susceptibilities to this parasite: BALB/c (susceptible), C57BL/6 (intermediate), and DBA/2 (resistant). Changes in ECM were observed mainly in BALB/c mice that, in general, presented tissue damage associated with high parasite burden. Under polarized light, Sirius Red revealed type I collagen that was predominant in the primary lesion in all strains studied at the early phase of infection, but gradually decreased and was replaced by abundant type III collagen fibres in chronic phase lesions. The presence of type III collagen seemed to provide support to inflammatory cells, mainly vacuolated and parasitized macrophages. Laminin expression was not altered during infection by L. (L.) amazonensis in any of the mouse strains studied. Furthermore, the decreased fibronectin expression, in all strains, in areas where amastigotes have been found, indicated that this decline was also not related to the genetic background.

Ectoparasitic copepods have been reported in a wide range of aquatic animals, including crustacean shellfish. However, with the exception of the salmon louse, Lepeophtheirus salmonis, our knowledge of such parasites in commercial species is rudimentary. The current study examines the morphology and pathology of the parasitic copepod, Nicothoë astaci (the ‘lobster louse’) in its host, the European lobster, Homarus gammarus. Lobsters were sampled from waters surrounding Lundy Island (Bristol Channel, UK) and all individuals collected were found to harbour female adult N. astaci in their gills, with a mean of 47·3 parasites/lobster. The majority of N. astaci were found in the basal region of pleurobranch gills. The parasite was found to attach to gill filaments via its oral sucker, maxillae and maxillipeds, and to feed on host haemolymph (blood) through a funnel-like feeding channel. It caused varying degrees of damage to the host gill, including occlusion of gill filaments and disruption to the vascular system in the central axis. Although there was evidence of extensive host response (haemocytic infiltration) to the parasite, it was displaced from the parasite attachment site and thus was observed in the central gill axis below. The region of gill filament immediately underlying the parasite feeding channel was devoid of such activity suggesting that the parasite interferes with the cellular defence and haemostatic mechanisms of the lobster in order to maintain invasion of the host.

The ultrastructure of penetrating stages of Babesia major merozoites responsible for causing trans–ovarian infection of the tick Haemaphysalis punctala was studied. These merozoites were different from forms of other Babesia species previously described in the tick in that they were highly pleomorphic. Most of the organelles found in these forms were similar to those found in other stages: however, the apical complex was found to be very active. Micronemes were found to the apical complex was found to be very active. Micronemes were found to be predominant in the cytoplasm of these merozoites and a cytoplasmic projection anterior to the polar ring and bounded by concentric rings was visible. A polar ring with radiating ribs, and microtubules was present. It is thought that the apical complex of these merozoites plays an important role in assisting the parasite to infect the ovary of the tick.

Cyclic GMP (cGMP) is responsible for various cellular functions including signal pathways and it acts as a mediator for nitric oxide (NO). In order to evaluate the anthelmintic efficacy of the plant-derived isoflavones, the crude peel extract of Flemingia vestita and pure genistein were tested with respect to the activity of nitric oxide synthase (NOS), NO efflux and the cGMP concentration in Rallietina echinobothrida, the cestode parasite of domestic fowl. For comparison, the parasites were also treated with genistein (the major isoflavone present in the crude peel extract), sodium nitroprusside (SNP), a known NO donor, and praziquantel (PZQ), the reference drug. At the time of onset of paralysis in the parasite, the activity of NOS showed a significant increase (35–46%) and a 2-fold increase of NO efflux into the incubation medium in the treated worms in comparison to the respective controls. The cGMP concentration in the parasite tissue increased by 46–84% in the treated test worms in comparison to the controls. The results show that the isoflavones, genistein in particular, from the crude peel extract of F. vestita influence the cGMP concentration in the parasite tissue, which plays a major role in the downstream signal pathways.

Hookworms are some of the most widespread of the soil-transmitted helminths (STH) with an estimated 438.9 million people infected. Until relatively recently Ancylostoma ceylanicum was regarded as a rare cause of hookworm infection in humans, with little public health relevance. However, recent advances in molecular diagnostics have revealed a much higher prevalence of this zoonotic hookworm than previously thought, particularly in Asia. This study examined the prevalence of STH and A. ceylanicum in the municipalities of Palapag and Laoang in the Philippines utilizing real-time polymerase chain reaction (PCR) on stool samples previously collected as part of a cross-sectional survey of schistosomiasis japonica. Prevalence of hookworm in humans was high with 52.8% (n = 228/432) individuals positive for any hookworm, 34.5% (n = 149/432) infected with Necator americanus, and 29.6% (n = 128/432) with Ancylostoma spp; of these, 34 were PCR-positive for A. ceylanicum. Considering dogs, 12 (n = 33) were PCR-positive for A. ceylanicum. This is the first study to utilize molecular diagnostics to identify A. ceylanicum in the Philippines with both humans and dogs infected. Control and elimination of this zoonotic hookworm will require a multifaceted approach including chemotherapy of humans, identification of animal reservoirs, improvements in health infrastructure, and health education to help prevent infection.

The adenophorean nematodes are evolutionarily distant from other species in the phylum Nematoda. Interspecific comparisons of predicted proteins have supported such an ancient divergence. Accordingly, Trichinella spiralis represents a basal nematode representative for genome sequencing focused on gaining a deeper insight into the evolutionary biology of nematodes. In addition, molecular characteristics that are conserved across the phylum could be of great value for control strategies with broad application. In this review, we describe and summarize progress that has been made on the sequencing and analysis of the T. spiralis genome. The genome sequence was used in preliminary analyses for the investigation of specific questions relating to the biology of T. spiralis and, more generally, to parasitic nematodes. For instance, we evaluated an unusually large DNase II-like protein family, predicted proteins of prospective interest in the parasite-host muscle cell interaction, anthelmintic targets and prospective intestinal genes, the encoded proteins (potentially) linked to immunological control against other nematodes. The results are discussed in relation to characteristics that are broadly conserved among evolutionary distant nematodes. The results lead to expectations that this genome sequence will contribute to advances in research on T. spiralis and other parasitic nematodes.

The three trypanosomatid genome projects have employed common strategies which include: analysis of pulsed-field gel electrophoretic chromosomal karyotypes; physical mapping using big DNA (cosmid, pacmid P1, bacterial artificial chromosome, yeast artificial chromosome) libraries; partial cDNA sequence analysis to develop sets of expressed sequence tags (ESTs) for gene discovery and use as markers in physical mapping; genomic sequencing; dissemination of information through development of web-sites and ACeDB-based fully integrated databases; and establishment of functional genomics programmes to maximize useful application of genome data. Highlights of the projects to date have been the demonstration that, despite extensive chromosomal size polymorphisms for diploid homologues within Africa trypanosomes, T. cruzi or Leishmania, the physical linkage groups for markers on each chromosome are retained across all isolates/species studied within each group. For African trypanosomes, detailed analysis of chromosome 1 has demonstrated that repetitive sequences and the two retroposon-like elements RIME and INGI are localized to a defined region at one end of the chromosome, with the bulk of the central region of the chromosome containing genes coding for expressed proteins. Comparative mapping shows that, although subtelomeric changes account for a large proportion of the polymorphism in chromosome size in African trypanosomes, there are significant expansions and contractions in regions across the entire chromosome. The highlight of the genomic sequencing projects has been the demonstration of just 2 putative transcriptional units of chromosome 1 of Leishmania major, extending on opposite strands from a point in the central region of the chromosome. A similar observation made on 93·4 kb of contiguous sequence for T. cruzi chromosome 3 suggests the presence of promoter and regulatory elements at the junctions of large polycistronic transcriptional units. All data obtained from the genome projects are made available through the public domain, which has prompted changing philosophies in how we approach analysis of the biology of these organisms, and strategies that we can employ now in the search for new therapies and vaccines.

Leucine aminopeptidases (LAPs) are a group of metalloexopeptidases that catalyse the sequential removal of amino acids from the N-termini of polypeptides or proteins. They play an important role in regulating the balance between catabolism and anabolism in living cells. LAPs of apicomplexa parasitic protozoa have been intensively investigated due to their crucial roles in parasite biology as well as their potentials as drug targets. In this study, we identified an M17 leucine aminopeptidase of Cryptosporidium parvum (CpLAP) and characterized the biochemical properties of the recombinant protein. Multiple sequence alignment of the deduced amino acid sequence of CpLAP with those of other organisms revealed that typical amino acid residues essential for metal binding and active-site formation in M17 LAPs were well conserved in CpLAP. Recombinant CpLAP shared similar biochemical properties such as optimal pH, stability at neutral pHs, and metal-binding characteristics with other characterized LAPs. The enzyme showed a marked preference for Leu and its activity was effectively inhibited by bestatin. These results collectively suggest that CpLAP is a typical member of the M17 LAP family and may play an important role in free amino acid regulation in the parasite.