Dirofilaria repens diagnosed by the presence of microfilariae in fine needle aspirates: a case report

BACKGROUND: Dirofilariasis due to Dirofilaria repens with viable microfilariae outside the worm have not been reported before. CASE: A 40-year-old truck driver from rural Shiraz, Iran, had a firm mass, 2.5 x 2.5 cm, at the dorsolateral aspect of the right forearm. Fine needle aspiration (FNA) was performed on 2 occasions. Several microfilariae with blunt heads, pointed posterior ends and empty caudal spaces resembling microfilariae of Wuchereria bancrofti but longer were seen. Since Iran is a nonendemic area for lymphatic filariae and the patient had a history of contact with a dog, with the impression of dirofilariasis, the mass was excised, and the presence of adult worms in tissue sections confirmed the diagnosis. CONCLUSION: This case ofsubcutaneous dirofilariasis was diagnosed by detecting microfilariae in FNA smears and was confirmed on histopathology.     

Isolation of microfilariae from blood by gravitational field-flow fractionation

Over 100 million persons suffer from diseases caused by filariae infestation, and one billion are at risk. A simple isolation method for both analytical and preparative separation is presented. Based on the simplest field-flow fractionation technique, the gravitational one, effective isolation of microfilariae is achieved. Microfilariae are eluted in the void volume of the channel without pollution by red blood cells. The red blood cell elution peak shows a total absence of microfilariae, as demonstrated after fraction collection and microscopic investigation. The elution mode of microfilariae and red blood cells appears to be a steric one, as confirmed by a reinjection experiment. The simplicity, low cost and the relatively short time required for this separation (10 min) indicate that gravitational field-flow fractionation could become a new separation tool for screening of microfilariae. With both live and dead microfilariae, the high recovery (66–80%) allows preparative fractionation for diagnostic purposes or fundamental research.     

Microfilariae in association with neoplastic lesions: report of five cases

Microfilariae and adult filarial worms have occasionally been detected in association with neoplastic lesions in cytological smears. The presence of microfilariae along with neoplasms is generally regarded as a chance association, yet some authors suggest that such parasitic infestations may be a causative factor for tumourigenesis. There are only a few reported cases in cytology literature documenting this association. We report the presence of microfilariae in routine cytology smears from one benign and four malignant tumours. Microfilariae could not be identified on histopathology available in four of these cases.     

Brugia malayi microfilaraemia in mice: a model for the study of the host response to microfilariae

Microfilariae of Brugia malayi were obtained from the peritoneal cavities of infected gerbils and were then injected intravenously into mice. A sub-periodic, nocturnal microfilaraemia was produced. The level of microfilaraemia was proportional to the number of parasites injected, with approximately 1-3% of microfilariae being found in the peripheral circulation. The duration of microfilaraemia was proportional to the number of parasites injected; it subsided by 30 days after injection of 104 microfilariae but was still present at a low level 120 days after injection of 2 x 105 microfilariae. A transient splenomegaly developed after injection of microfilariae. Histopathological examination revealed large numbers of microfilariae free in the lumens of pulmonary small blood vessels and without any accompanying inflammatory reaction. Lesser numbers of microfilariae were seen in the cardiac blood and hepatic and renal blood vessels for the first few days after injection. There was cellular proliferation in the splenic white pulp and vascular congestion of the red pulp. Microfilariae labelled with 51Cr were injected intravenously; 57% of radioactivity was found in the lungs, 8.5% in the liver and 2.9% in the spleen. Mice developed immediate hypersensitivity reactions to B. malayi antigen by 4 weeks after injection, but Arthus and delayed hypersensitivity reactions were not seen at any time. when mice which had been injected 5 months previously were challenged with a 2nd injection of microfilariae, there was an accelerated clearance of parasites over 2 weeks and a marked peripheral blood eosinophilia developed. In contrast with natural infections, in which the continuous production of microfilariae complicates assessment, this model provides a system in which factors controlling the circulation of microfilariae in the bloodstream can be studied independently.     

Effect of N-acetyl-D-glucosamine on the migration of Brugia pahangi microfilariae into the haemocoel of Aedes aegypti

Two strains of Aedes aegypti (L.), differing in their susceptibility to Brugia pahangi (Buckley & Edeson), were examined with regard to the effect on the proportion of microfilariae migrating from the mid-gut, of specific carbohydrate supplements in the infecting bloodmeal. N-Acetyl-D-glucosamine (GlcNAc), a sugar also present on the microfilarial sheath, significantly increased the migration rate. This enhancement is greater for the refractory strain of Ae.aegypti. The use of sucrose as a control sugar results in no enhancement of microfilariae migration. It is postulated that the GlcNAc is acting by blocking endogenous gut/peritrophic membrane carbohydrate binding proteins, which would normally inhibit microfilariae migration. Furthermore, there is a significant correlation whereby increasing loads of microfilariae ingested result in decreasing proportions migrating across the mid-gut.     

Efficacy of moxidectin for the prevention of adult heartworm (Dirofilaria immitis) infection in dogs

The authors report the efficacy of orally administered moxidectin for the prevention of canine heartworm infection in two endemic areas in northern Italy. Two trials were conducted on a total of 257 dogs, including 137 treated with moxidectin (minimum dose of 3 mcg/kg body weight), 85 with ivermectin (minimum dose 6.6 mcg/kg b.w.) and 35 untreated controls. Results of testing for microfilariae and circulating adult female antigens were negative for all treated dogs at the end of both trials. No adverse reactions to moxidectin were observed. In the study areas, prevalence values for Dirofilaria immitis infection calculated on the basis of the untreated controls and testing dogs which had no preventive treatment in the previous transmission season ranged 23-65%. This study confirms the efficacy and safety of moxidectin in the prevention of adult heartworm infection in dogs.     

Immunological tolerance: The key feature in human filariasis?

Filariasis is a widespread tropical disease caused by a group of nematode parasites that can survive for many years in immunocompetent hosts. The paradox of filariasis has always been the inverse association between parasite density, in terms of circulating microfilariae in the blood, and severe pathology. In this review, Rick Maizels and Rachel Lawrence argue that microfilariae and adult parasites induce a form of immunological tolerance which prevents both parasite elimination and progression to disease. The breakdown of this unresponsiveness is seen as the critical step towards pathogenesis. However, not every exposed individual progresses through infection to disease. The authors discuss evidence for protective immunity acting on antigens from the mosquito-borne infective larva, and propose that this stage represents a vulnerable target outside the scope of tolerance and pathogenesis. Stage-specific larval antigens, to which asymptomatic hosts are known to respond, may therefore represent the most effective and safe choice for an anti filarial vaccine.     

Lesions in the liver and kidney of Dirofilaria immitis-infected dogs following treatment with ivermectin

Six dogs with spontaneous heartworm disease were injected with a single dose of ivermectin. After 48 h of treatment, microfilariae counts were reduced by 92%-98% of pretreatment counts. In pretreatment biopsies examined by light and electron microscopy, microfilariae were unaltered in the sinusoids of the liver and also in the glomerular capillaries and interstitial blood vessels of the kidney. However, there was irregular thickening and dense deposits in the basement membranes of glomerular capillaries, along with a modest increase in mesangial cells and matrix. In post-treatment liver biopsies examined by light microscopy, there were numerous granulomas in the sinusoids which contained degenerated microfilariae. In post-treatment kidney biopsies there was moderate thickening of glomerular basement membranes along with pronounced proliferation of mesangial cells and matrix. Glomerular capillaries were partially or completely occluded by degenerated microfilariae. In addition, there were interstitial granulomas in the kidney. It was observed with the aid of electron microscopy that highly vacuolated and degenerated microfilariae were incorporated into granulomas in the liver sinusoids of post-treatment biopsies. In post-treatment kidney biopsies glomerular capillaries were usually occluded by degenerated microfilariae. Basement membranes were thickened and contained dense deposits. Mesangial cells and matrix were extensively increased. Interstitial granulomas in the kidney contained dead microfilariae.     

Kinetic Memory Based on the Enzyme-Limited Competition

Cellular memory, which allows cells to retain information from their environment, is important for a variety of cellular functions, such as adaptation to external stimuli, cell differentiation, and synaptic plasticity. Although posttranslational modifications have received much attention as a source of cellular memory, the mechanisms directing such alterations have not been fully uncovered. It may be possible to embed memory in multiple stable states in dynamical systems governing modifications. However, several experiments on modifications of proteins suggest long-term relaxation depending on experienced external conditions, without explicit switches over multi-stable states. As an alternative to a multistability memory scheme, we propose “kinetic memory” for epigenetic cellular memory, in which memory is stored as a slow-relaxation process far from a stable fixed state. Information from previous environmental exposure is retained as the long-term maintenance of a cellular state, rather than switches over fixed states. To demonstrate this kinetic memory, we study several models in which multimeric proteins undergo catalytic modifications (e.g., phosphorylation and methylation), and find that a slow relaxation process of the modification state, logarithmic in time, appears when the concentration of a catalyst (enzyme) involved in the modification reactions is lower than that of the substrates. Sharp transitions from a normal fast-relaxation phase into this slow-relaxation phase are revealed, and explained by enzyme-limited competition among modification reactions. The slow-relaxation process is confirmed by simulations of several models of catalytic reactions of protein modifications, and it enables the memorization of external stimuli, as its time course depends crucially on the history of the stimuli. This kinetic memory provides novel insight into a broad class of cellular memory and functions. In particular, applications for long-term potentiation are discussed, including dynamic modifications of calcium-calmodulin kinase II and cAMP-response element-binding protein essential for synaptic plasticity.     

Brugia malayi: clearance of microfilaremia induced by diethylcarbamazine independently of antibody

The microfilaricidal effect of diethylcarbamazine was studied in the murine model of Brugia malayi microfilaremia. CFI mice with circulating microfilariae were treated with either diethylcarbamazine (6 mg kg-1 day-1) for 10 days or with normal saline. Although antibodies against crude microfilarial antigen were detected in both groups at the time of completion of therapy, the chemically induced microfilarial clearance occurred prior to the development of these antibodies. In addition, the passive transfer of antibody just prior to chemical treatment did not enhance diethylcarbamizine-induced microfilarial clearance. In vitro studies confirmed previous observations that diethylcarbamazine enhanced antibody-dependent neutrophil adherence. Microfilarial clearance was found to be due to trapping and lysis in the liver on pathologic examination. These studies demonstrate that diethylcarbamazine leads to microfilarial clearance in the liver by mechanisms that are not dependent on host antibody.     

In vitro study on humoral encapsulation of microfilariae: effects of diethyldithiocarbamate and dopachrome on the reaction

The effects of a phenoloxidase inhibitor, diethyldithiocarbamate, and an intermediate of the melanin biosynthetic pathway, dopachrome, on the humoral encapsulation of microfilariae were studied in vitro. Diethyldithiocarbamate inhibited the melanization but did not prevent the humoral encapsulation of microfilariae. In the presence of diethyldithiocarbamate in the in vitro system, transparent material in the form of numerous granules were deposited on the surface of the microfilariae and coalesced to form a consolidated transparent capsule around microfilariae. The thickness of transparent capsule was significantly greater than that of normal melanotic capsule. The transparent capsule material, which was probably the precursor of the melanotic capsule, was demonstrated histochemically as a protein-carbohydrate complex. The ultrastructure of transparent capsule showed that the transparent capsule material was very different from that of melanotic capsule material, being more flocculent and less granular in appearance. When dopachrome was concurrently present with the inhibitor in the in vitro encapsulation system, it failed to restore normal melanotic encapsulation. The data presented lead to the following conclusions: 1. The melanin biosynthetic pathway in the humoral encapsulation of microfilariae is similar to that found in the synthesis of mammalian melanin. 2. The humoral encapsulation of the microfilariae in vitro is a two-step process. The protein-carbohydrate complex is deposited on the surface of microfilariae. Then the tyrosine group of the complex is catalysed by phenoloxidase to melanin.     

Role of fine needle aspiration cytology in detection of microfilariae: report of 2 cases

BACKGROUND: Filariasis is a major public health problem in developing countries, and the diagnosis is conventionally made by demonstrating microfilariae in the peripheral blood smear. However, microfilariae have been incidentally detected in fine needle aspirates of various lesions in clinically unsuspected cases of filariasis with absence of microfilariae in the peripheral blood. CASES: In case 1, a 21-year-old woman presented with multiple left axillary lymphadenopathy of 3 months' duration. In case 2, a 32-year-old woman presented with a thyroid nodule of 7 months' duration. Fine needle aspiration smears from both cases showed sheathed microfilariae of Wuchereria bancrofti. In both cases, microfilariae could not be demonstrated in the peripheral blood smears and the blood eosinophil counts were within normal limits. The histopathologic examination showed neither microfilariae nor adult worm. CONCLUSION: Although microfilariae in cytologic material are considered incidental findings, these cases illustrate the value of routine fine needle aspiration cytology in the detection of asymptomatic and clinically unsuspected cases of bancroftian filariasis. Absence of microfilariae in the peripheral blood does not exdude filarial infection.     

周期型马来丝虫微丝蚴寿命的观察

将实验感染周期型马来丝虫的长爪沙鼠的微丝蚴蚴阳性腹腔稀释液,移注于正常沙鼠腹腔内,微丝蚴除能在腹腔内长期生存外,还可出现于外周血液中,其在外周血液内末次阳性检出时间最长可超过32周,在腹腔液内末次阳性检出时间最长为77周,故马来微丝蚴在沙鼠外周血液中的最长寿命不短于7.5月,而在腹腔液内的最长寿命可超过1.5年以上。     

Cytologic detection of Wuchereria bancrofti microfilariae in urine collected during a routine workup for hematuria

A 23-year-old Guyanese man experienced intermittent, total, painless, gross hematuria for a month for which he sought medical attention at the Kings County Hospital Center in Brooklyn. Hematuria was accompanied by weakness but not by frequency of urination or burning on urination. Catheterized urine at the time of cystoscopy and each of two subsequent voided specimens examined cytologically contained sheathed microfilariae. Distinguishing features of the microfilariae were well demonstrated with the Papanicolaou stain. The well-stained nuclei, which did not extend into the clear zone, and the distinct, pale-stained sheath led to the positive identification of the microfilariae as Wuchereria bancrofti. The Papanicolaou stain may well be the stain of choice for the identification of microfilariae in the blood. The excellent detail obtained with this routine cytologic stain is as good as that with Giemsa, which does not stain the sheath.     

Microfilarial perforation of the midgut of a mosquito

To determine whether the midgut envelope of mosquitoes is disrupted by the passage of microfilariae, ultrastructural changes induced by microfilariae of Brugia malayi were observed in midguts of Aedes aegypti mosquitoes. Basal and apical plasma membranes were destroyed, disrupting the full depth of the midgut wall. Ingested ferritin lay against the gut wall, suggesting absence of the peritrophic membrane during penetration. Exsheathment of microfilariae appears to be enhanced by movement against the constricting midgut wall. It was concluded that particles present in the lumen of the gut may be disseminated passively to the hemocoel.     

Microfilariae in association with other diseases. A report of six cases

BACKGROUND: Lymphatic filariasis is a major public health problem in tropical countries. Earlier reports have reported microfilariae as an incidental finding in body fluids and fine needle aspiration smears from various sites. CASES: The findings of body fluid cytology and fine needle aspiration smears from six patients with microfilariae in association with other conditions--tubercular pleural effusion/lymphadenitis, pregnancy and non-Hodgkin's lymphoma--are presented. Three patients demonstrated an associated eosinophilic cellular exudate. Adherence of inflammatory cells to microfilariae was seen in two patients. CONCLUSION: Although microfilariae in cytologic smears are considered incidental findings, the association of microfilariae with debilitating conditions suggests that it is an opportunistic infection and needs further study.     

Brugia malayi: intravenous injection of microfilariae in ferrets as an experimental method for occult filariasis

Microfilaremia, immune responses, and pathology were compared in ferrets infected with 100 third-stage larvae of Brugia malayi (subperiodic strain) or injected intravenously with 10(6) microfilariae. Ferrets (Mustela putorius furo) inoculated with third-stage larvae typically became patent during the third month after infection, with a mean patency of 123 +/- 25 (SE) days. Ferrets injected intravenously with microfilariae exhibited a relatively constant microfilaremia for 3-4 weeks and usually cleared microfilariae before the fourth month. Ferrets that cleared microfilariae after intravenous injection of microfilariae or after infection with third-stage larvae failed to become patent or became amicrofilaremic within 3 weeks after a challenge intravenous injection of 10(6) microfilariae. Clearance of circulating microfilariae was associated with eosinophilia and serum antibody specific for the microfilarial sheath in ferrets injected with microfilariae and in most ferrets infected with third-stage larvae. Ferrets infected with third-stage larvae and necropsied after clearance of microfilariae had tissue inflammatory reactions to microfilariae characteristic of occult filariasis (tropical eosinophilia) in man; these ferrets exhibited immediate cutaneous hypersensitivity and circulating reaginic antibody to antigens of microfilariae. In ferrets necropsied following two intravenous injections of microfilariae, the majority of ferrets examined within 10 days after clearance of microfilariae had visible liver lesions to microfilariae identical to those of the ferrets infected with third-stage larvae; immediate cutaneous hypersensitivity and reaginic antibody were not consistently detected in ferrets injected with microfilariae. Sera from ferrets that had cleared circulating microfilariae were transferred passively into ferrets made microfilaremic by intravenous injection of microfilariae. Sera with microfilarial sheath-reactive IgG antibody titers (greater than or equal to 1:200) and microfilarial agglutination titers (greater than or equal to 1:40) rapidly cleared injected microfilariae (less than 24 hr); this serum also cleared or greatly reduced circulating microfilariae established by an infection with third-stage larvae; only the IgG-containing fraction of the sera was active in immune clearance. Sera that cleared microfilariae of B. malayi did not clear circulating microfilariae of Dirofilaria immitis or prevent recurrence of circulating microfilariae of B. malayi in ferrets infected with adult filariae.(ABSTRACT TRUNCATED AT 400 WORDS)     

An ultrastructural study on the encapsulation of microfilariae of Brugia pahangi in the haemocoel of Anopheles quadrimaculatus

Chen C. C. and Laurence B. R. 1985. An ultrastructural study on the encapsulation of microfilariae of Brugia pahangi in the haemocoel of Anopheles quadrimaculatus. International Journal for Parasitology15: 421–428. The encapsulation of microfilariae of Brugia pahangi in the haemocoel of Anopheles quadrimaculatus was studied ultrastructurally. The microfilariae was first seen enclosed in an acellular electron dense capsule as early as 10 min after the engorgement of the mosquitoes from a cat parasitized by filariae. Two hours later, the mosquito plasmatocytes spread onto and around the humoral capsule. A completed capsule, which was seen at 24–48 h, was composed of an inner humoral layer and outer cellular layer. After 1 week, some electron dense haemocytes were seen attached to the outer surface of the cellular layer. These results suggested that the encapsulation of microfilariae in the haemocoel of mosquitoes combines both humoral and cellular reaction; humoral encapsulation occurs first and cellular encapsulation takes place later. The significance of combined reactions of humoral and cellular encapsulation in the mosquito-filarial system is discussed with reference to the encapsulation reaction of other insects.