Wolbachia spp. interfere na detecção molecular de Ehrlichia canis e Anaplasma platys em cães com microfilaremia / Wolbachia spp. interferes with molecular detection of Ehrlichia canis and Anaplasma platys in microfilaremic dogs

This report aimed to study the interference in molecular testing for Ehrlichia canis and Anaplasma platys in blood of 155 dogs from the coastal region of Rio de Janeiro. Five Anaplasmataceae positive samples but negative for E. canis and A. platys, from microfilaremic animals, were chosen for sequencing. These sequences, when compared to Gen et Bank database, showed 88% to 100% similarity with Wolbachia spp. denoting an interference in the detection of DNA from other members of Anaplasmataceae, possibly due to a high concentration of Wolbachia spp. DNA.(AU)

Detection of Wuchereria bancrofti DNA in paired serum and urine samples using polymerase chain reaction-based systems

The Global Program for the Elimination of Lymphatic Filariasis (GPELF) aims to eliminate this disease by the year 2020. However, the development of more specific and sensitive tests is important for the success of the GPELF. The present study aimed to standardise polymerase chain reaction (PCR)-based systems for the diagnosis of filariasis in serum and urine. Twenty paired biological urine and serum samples from individuals already known to be positive for Wuchereria bancrofti were collected during the day. Conventional PCR and semi-nested PCR assays were optimised. The detection limit of the technique for purified W. bancrofti DNA extracted from adult worms was 10 fg for the internal systems (WbF/Wb2) and 0.1 fg by using semi-nested PCR. The specificity of the primers was confirmed experimentally by amplification of 1 ng of purified genomic DNA from other species of parasites. Evaluation of the paired urine and serum samples by the semi-nested PCR technique indicated only two of the 20 tested individuals were positive, whereas the simple internal PCR system (WbF/Wb2), which has highly promising performance, revealed that all the patients were positive using both samples. This study successfully demonstrated the possibility of using the PCR technique on urine for the diagnosis of W. bancrofti infection.

Avaliação da acurácia de diferentes testes laboratoriais no diagnóstico da filariose em crianças e adolescentes / Evaluation of the accuracy of different laboratory tests for the diagnosis of filariasis in children and adolescents

Apesar de a filariose linfática (FL) ser considerada uma doença de adultos, estima-se que 22 milhões de crianças 15 anos de idade estejam infectadas no mundo. A Organização Mundial da Saúde tem recomendado o monitoramento da infecção nessa população. Este estudo teve como objetivo comparar a acurácia dos métodos parasitológicos e imunológicos no diagnóstico da filariose bancroftiana em escolares. O estudo foi desenvolvido em escolares, com idade entre 4 e 15 anos, residentes em três bairros do município de Olinda - Pernambuco, Brasil. As amostras de sangue capilar e venoso foram coletadas entre 23:00 e 1:00 hora da manhã. Em seguida, as amostras de sangue venoso foram guardadas para posterior realização das técnicas de filtração, concentração de Knott, Og4C3 - ELISA. Amostras de sangue capilar foram obtidas para confecção das lâminas de gota espessa e realização do teste rápido de imunocromatografia (ICT). As médias de densidade de microfilaremia foram calculadas em escala logarítmica. A acurácia dos testes foi avaliada em relação ao padrão-ouro (filtração). A especificidade do Og4C3 e ICT foi estimada utilizando a equação de Staquet et al. (1981). Um total de 805 escolares foi examinado. As médias de antigenemia filarial foram mais elevadas entre as crianças residentes nos bairros de alto da conquista e alto da bondade. As cargas parasitárias e os níveis de antigenemia não variaram com idade e sexo. A prevalência de microfilaremia pela técnica de filtração, de 5,2 por cento, foi a mais elevada em relação às demais técnicas parasitológicas. A prevalência de antigenemia filarial pelo teste Og4C3 foi de 17,4 por cento. Na comparação da acurácia dos testes em relação ao padrão-ouro, as técnicas de gota espessa e Knott apresentaram valores de sensibilidade de 85,2 por cento, inferiores aos testes ICT e Og4C3, que foi de 100 por cento. Conclui-se que as técnicas de filtração e Og4C3 são as mais apropriadas para a avaliação de transmissão em áreas com programas de eliminação em andamento.

Ocorrência da Mansonella ozzardi (Nematoda, Onchocercidae) em comunidades ribeirinhas do rio Purus, Município de Boca do Acre, Amazonas, Brasil / Occurrence of Mansonella ozzardi (Nematoda, Onchocercidae) in riverine communities of the Purus river, Boca do Acre municipality, Amazonas State, Brazil

Este estudo teve o objetivo de estimar as prevalências de Mansonella ozzardi e calcular taxa de infecção parasitária nos simulídeos. O trabalho foi realizado em comunidades ribeirinhas do rio Purus, Boca do Acre, Amazonas, Brasil. As prevalências foram obtidas por meio do método de gota espessa de sangue obtido por porção digital. Os simulídeos coletados foram dissecados para estimar a taxa de infecção parasitária. A prevalência de M. ozzardi foi de 27,3 por cento (282/77). Foram observadas maiores prevalências nos homens (31,97 por cento) que nas mulheres (22,22 por cento), nos agricultores (48 por cento) e nos indivíduos entre: 38-47 (60 por cento), 48-57 (66,66 por cento) e 58-67 (75 por cento). A microfilaremia foi maior nos indivíduos entre 58-67 anos (média = 58,41mf/40µL), sexo masculino (41,44mf/40µL) e nos agricultores (49,94mf/40µL). Somente o simulídeo Cerqueirellum amazonicum foi encontrado infectado com taxa de infecção parasitária de 0,98 por cento.

The study aimed to estimate the prevalence of Mansonella ozzardi and calculate the parasitic infection rate in simuliid blackflies. The research was conducted in communities on the Purus River, Boca do Acre municipality, Amazonas State, Brazil. Prevalence was measured using the thick smear method. Captured blackflies were dissected to verify the parasitic infection rate. M. ozzardi prevalence was 27.30 percent (77/282). The study showed higher prevalence in men (31.97 percent) than women (22.22 percent), farmers (48.99 percent), and individuals in the 38-47 (60.00 percent), 48-57 (66.66 percent), and 58-67-year age brackets (75.00 percent). Microfilaremia was higher in individuals 58 to 67 years of age (average= 58.41mf/40µl), men (41.44mf/40µl), and farmers (49.94mf/40µl). Only the simuliid Cerqueirellum amazonicum was found infected with a parasitic infection rate of 0.98 percent.

Acometimento ocular em pacientes com mansonelose / Ocular manifestations in mansonelliasis

OBJETIVOS: Avaliar as alterações oculares observadas em indivíduos com mansonelose residentes na área rural do município de Coari, AM, na região do médio Amazonas. MÉTODOS: Foi conduzido um estudo de corte transversal envolvendo 543 moradores residentes na área rural do município de Coari, na margem direita do rio Solimões. Todos os indivíduos foram submetidos a exame de gota espessa para pesquisa de microfilária e os indivíduos positivos foram submetidos a exame oftalmológico. Lesões oculares suspeitas foram biopsiadas e enviadas para exame histopatológico. RESULTADOS: Mansonella ozzardi foi encontrada no sangue periférico em 103 (18,9 por cento) indivíduos. Noventa e cinco pacientes com mansonelose foram examinados, observando ceratite puntata em 12 (11,7 por cento) destes, ceratite numular em um indivíduo (1 por cento) e esclerosante em outro (1 por cento). Biópsia de pele foi negativa em todos os indivíduos. Biópsia de conjuntiva e limbo foi realizada em cinco pacientes com mansonelose e biópsia de córnea em outros três pacientes. Duas biópsias de conjuntiva e limbo e uma biópsia de córnea foram enviadas para exame histopatológico, não sendo observadas microfilárias ou alterações sugestivas da presença destas nas peças estudadas. O restante das biópsias foi enviado para pesquisa de DNA do parasita por PCR, não se encontrando microfilária. CONCLUSÕES: Associação entre infecção por Mansonella ozzardi e lesões corneanas sem etiologia aparente foi verificada neste estudo, mas a ausência de achados específicos no exame histopatológico e a não identificação de microfilárias por PCR em lesões suspeitas põem em dúvida a o acometimento corneano na mansonelose. Estudos mais amplos de populações com alta prevalência de M. ozzardi e a realização de outros testes para identificação de microfilárias em tecido ocular são necessários para investigar a patogenicidade da microfilária ao tecido ocular.

PURPOSE: To evaluate eye manifestations in mansonelliasis patients from Coari, AM, rural area on the middle Amazonas River. METHODS: Cross-sectional study consisted of evaluation of 543 subjects from Coari rural area on Solimões River right margin. All subjects were submitted to blood test for microfilaria and the subjects who had a positive test also were submitted to skin biopsies and ophthalmologic examination. Histopathology and PCR research for microfilaria were done on eye biopsies of suspect lesions. RESULTS: Mansonella ozzardi was found in peripheral blood of 103 (18.9 percent) subjects. Ninety-five mansonelliasis patients were examined. Punctate keratitis was observed in 12 of them, nummular keratitis in one subject and sclerosing keratitis in another one. Skin biopsies were negative in all subjects. Conjunctival and limbal biopsies were done in five mansonelliasis patients and corneal biopsy in another three patients. Conjunctival and limbal biopsies of two patients and corneal biopsy of one patient showed no microfilaria or alterations due to its presence on histopathology. PCR showed no evidence of microfilaria in the other biopsies. CONCLUSION: Association between Mansonella ozzardi infection and corneal lesions with no evident etiology was observed in this study, but the lack of conclusive findings on histopathology and PCR make us doubt the corneal involvement in mansonelliasis. Large studies of high mansonelliasis prevalence populations and extensive employment of microfilaria identification tests in ocular tissue are needed to evaluate the microfilaria corneal pathogenicity.

Microfilaria in thyroid gland nodule.

Presence of microfilaria in the thyroid gland is a rare finding. Filariasis is a common public health problem in the Indian sub-continent. Most of the cases of microfilaria in thyroid gland reported in the literature are associated with goiter and thyroid neoplasms. Here, we present a rare case that showed microfilaria on fine needle aspiration cytology of solitary thyroid nodule.

Kinetics of microfilaraemia & antigenaemia status by Og(4)C(3) ELISA in bancroftian filariasis.

BACKGROUND & OBJECTIVE: Bancroftian filariasis caused by Wuchereria bancrofti is endemic in many parts of India. In recent years diagnosis of W. bancrofti infection has been revolutionized with the availability of filarial antigen tests, which is important in monitoring success of chemotherapy. We carried out this study to measure microfilariaemia and antigenemia levels in bancroftian microfilariae (mf) carriers at 1 yr follow up after chemotherapy, in lymphoedema patients and in endemic controls from a filariasis endemic area in Tamil Nadu State using Og(4)C(3) ELISA to identify the best marker to assess success of chemotherapy. METHODS: Serum samples were collected from 30 bancroftian microfilaremic (Mf) carriers pre-treatment and at sequential intervals (7,30,60,90,180 and 365 days) following treatment with diethylcarbamazine (DEC:6mg/kg body weight, single dose), 30 lymphoedema patients (without treatment) at periodic intervals, and 68 control subjects (24 endemic normal subjects in filariasis endemic area in Tamil Nadu State, 24 non-endemic normal subjects residing in Chandigarh, India; 5 brugian filariasis, 5 endemic control subject in brugian filariasis endemic area and 10 other disease controls). The circulating antigen of W. bancrofti was measured quantitatively using Og(4)C(3) ELISA kit. RESULTS: In Mf carriers, there was no significant difference in microfilariae count in pre- and post-treatment (PT) samples till day 30 while significant differences were observed in pre- and sequentially collected post-treatment (PT) samples day 60 to 180 (P<0.001), day 365 (P<0.005). However, there was no significant difference in antigenaemia levels between pre-treatment (day 0) and PT samples collected on day 7 onwards till day 365. Though of the 19 patients who could be followed up till 365 days PT, 4 (21%) were amicrofilaraemic, none became antigen negative. No significant difference was found in antigenaemia levels in sequentially collected samples from lymphoedema patients. Significant differences were observed in antigenaemia levels in samples collected at the start of study in mf carriers as compared to lymphoedema patients and endemic normal subjects (P<0.001). Subjects (non-endemic control) residing in filariasis free area (24), brugian endemic area (5), B.malayi infected patients (5) and patients with other parasitic diseases (10) were found antigen negative. INTERPRETATION & CONCLUSION: Annual single dose of DEC therapy alone may not result in complete clearance of infection and detection of antigenaemia rather than microfilaraemia may be taken into consideration as an indicator of successful chemotherapy. The study supports the earlier view that filarial antigenaemia is relatively common in amicrofilaraemic and asymptomatic subjects in endemic areas and further studies are needed to determine the clinical significance, prognosis and effective management of such infections in endemic areas.

Microfilariae in bone marrow aspiration smears, correlation with marrow hypoplasia: a report of six cases.

Microfilariae have been reported in various cytological preparations, however there are very few case reports describing microfilariae in bone marrow aspirates. We report six such cases with the interesting finding of marrow hypoplasia in five of these cases.

Filariasis in Ghatampur Tahsil of Kanpur Nagar District: indications of high endemicity locus.

A study in the 26 villages surveyed, the mf rate was observed to vary from 6.4% to 17.8%, the disease rate ranged from 1.9% to 10% and total infection rate from 8.2% to 26.4%. The median microfilaraemia density among positives was 10 and 90% of persons had density below 60 and in 10% above this level. The mf rate among those who never used bednets while sleeping was found to be 11.8%, 15.7% higher than 10.2% among those who ever used bednets (8.7% in regular users and 10.7% among irregular users) to protect from mosquitoes bites (p < 0.05). The lymphatic disease was found to be 3.8%; 3.7% in males and 4.1% in females. Of the males, 16.3% had acute disease, 51.8%) hydroceles of varying grades and 32% edema of different grades including elephantiasis. Of the females with lymphatic disease, 25.6% acute disease, 62.8% edema including elephantiasis and 11.6% had mastitis. The study indicates that area is endemic for filariasis and needs control programmes.

A pilot study on the status of lymphatic filariasis in a rural community of Bihar.

A pilot study of lymphatic filariasis was conducted in two contiguous villages of Patna district in Bihar situated at the side of the river Ganges, known to be endemic for lymphatic filariasis, to study present status of transmission parameters of filariasis. Of the 1872 persons examined, 8.4% were found asymptomatic but microfilaraemic. Morbidity pattern due to filarial infection showed an increase with advancement of age and significantly high in males as compared to female (p < 0.001). Acute and chronic filarial disease was observed as 0.5% and 9% respectively. Microfilaria was found in 10% of acute and 11.2% of chronic filarial cases. The Mf rate was found to be 9.9% in males and 9.0% in females respectively. The parasite species was identified as W. bancrofti. The vector fauna surveyed show highest prevalence of vector species of Cx. quinquefasciatus (43%) in both domestic as well as predomestic area in the community. Other species like Cx. vishnui and Ma. uniformis were also seen. Each household and predomestic area was searched for mosquito fauna at night. The infection rate in vectors was found to be 14% and infectivity rate (L3) was 8%. The filariasis cases detected in the study were treated with 12 days course of DEC 6 mg/kg body weight.

Lymphatic filariasis in rural areas of Patna District, Bihar. A challenge ahead.

Patna district was endemic for lymphatic filariasis (LF). During November 2004, a lymphatic filariasis survey was carried out in seven randomly selected villages from four PHCs of Patna district. Of 1878 night blood smears (NBS) examined, 117 were found positive for W. bancrofti infection (mf rate 6.2%). Microfilaria carriers were detected from all surveyed villages. In all areas prevalence of microfilaria generally increased with age to maximum 15-34 years and then decline within most age-groups. More males (6.4%) were affected than the females (5.8%). All microfilaria (mf) carriers were residents of Patna district. Over all disease rate was 9.1%. Out of 171 diseased individuals, 121 persons were having hydrocele (6.4%). Disease rate was higher in males (12.0%) than females (4.1%). The mean mf density was 11.7. There was no significant difference of mean mf density in males and females. Vector density ranged from 205 to 780 per ten man hours. The survey indicates that the filariasis situation remained unchanged since last fifty years and still is one of the major public health problem in surveyed areas.

A baseline study of rural Bancroftian filariasis in southern India.

Night mass blood surveys were carried out for parasitological evidence of Bancroftian filariasis in 45 rural areas belonging to 9 National Filaria Control Program (NFCP) zones of East Godavari and West Godavari districts of Andhra Pradesh, India during the period 1998 to 2001. Mf prevalence range between 2.9 to 10.2%, and mf intensities in 20 mm3 blood samples ranged from 1-281. The present study explains the trend of microfilaria dynamics in the rural population, where mass drug delivery has been implemented since 1997, and anti-larivicidal and adulticidal control measures have not been adopted.

The preferential site of adult Wuchereria bancrofti: an ultrasound study of male asymptomatic microfilaria carriers in Pondicherry, India.

BACKGROUND: The traditional method of detection of microfilaria in night blood specimens for the diagnosis of Wuchereria bancrofti infection is being replaced with circulating filarial antigen in day blood specimens, which has a high sensitivity. However, both methods are indirect tests to detect the presence of adult worms in vivo. Localization of adult worms in vivo in their natural habitat may help in understanding better the end-point of drug treatment, the adulticidal action of antifilarial drugs, and in locating the site of lymphatic pathology. We used ultrasound examination to assess the preferential location of adult worms in an area endemic for lymphatic filariasis. METHODS: Ultrasound examination was done in 36 asymptomatic male carriers of Wuchereria bancrofti microfilaria to detect the location of adult worms. Both sides of the scrotum (root of the scrotum, epididymis, spermatic cord, testis and the adjoining area), lymphatic vessels and inguinal, popliteal, axillary and epitrochlear lymph nodes were examined using a 7.5 MHz probe in real-time B mode. RESULTS: The 'filaria dance sign (FDS)' suggesting the presence of adult worms was observed in 22 carriers (61%). The preferential site of location of the adult worms was the intrascrotal juxtatesticular lymphatic vessels in 'nests' along the lymphatic vessels of the epididymis, spermatic cord and paratesticular region. The number of nests varied between 1 and 4 with a mean size of 0.3 cm2. In 95% of cases, localization of the worms was unilateral. The mean microfilaria (SD) count-positive cases for those with the filarial dance sign (264 [199]) was significantly higher (p<0.05) than for the negative cases (171 [196]). CONCLUSION: Ultrasound visualization of adult worms of Wuchereria bancrofti in vivo is possible and confirms the concept that the worms have their own territory and reside in 'nests'. The preferential site of localization of the adult worms in men is the intrascrotal juxtatesticular lymphatic vessels.

Eye lesions and onchocerciasis in a rural farm settlement in Delta State, Nigeria.

Between December 1999 and September 2000, an investigation on 326 inhabitants who voluntarily participated in screening for eye lesions and onchocerciasis in a rural farm settlement in Aniocha North local government area of Delta State, Nigeria was carried out. Microfilariae were reported in the skin tissues of 134 (41.1%) inhabitants. Of these, males had more microfiladermia (49.9%) than their female counterparts (33.3%). The percentage prevalence of the clinical signs of onchocerciasis in the farm settlement was hanging groin 14.1%, onchocercal nodules 25.1%, onchocercal dermatitis 27.6% and scrotal enlargement 3.7%. The percentage prevalence of the eye lesions among the onchocerciasis infected inhabitants was cataract 7.4%, glaucoma 3.7%, optic atrophy 0.6% and uveitis 18 (5.5%). Impaired vision occurred among 80 (24.5%) inhabitants and blindness was reported among 2 (0.6%) inhabitants.

Bloodmeal microfilariae density and the uptake and establishment of Wuchereria bancrofti infections in Culex quinquefasciatus and Aedes aegypti

The relationship between ingestion of microfilariae (mf), production of infective larvae (L3) and mf density in human blood has been suggested as an important determinant in the transmission dynamics of lymphatic filariasis. Here we assess the role of these factors in determining the competence of a natural vector Culex quinquefasciatus and a non vector Aedes aegypti to transmit Wuchereria bancrofti. Mosquitoes were infected via a membrane feeding procedure. Both mosquito species ingested more than the expected number of microfilariae (concentrating factor was 1.28 and 1.81 for Cx. quinquefasciatus and Ae. aegypti, respectively) but Cx. quinquefasciatus ingested around twice as many mf as Ae. aegypti because its larger blood meal size. Ae. aegypti showed a faster mf migration capacity compared to Cx. quinquefasciatus but did not allow parasite maturation under our experimental conditions. Similar proportions of melanized parasites were observed in Ae. aegypti (2.4 per cent) and Cx. quinquefasciatus (2.1). However, no relationship between rate of infection and melanization was observed. We conclude that in these conditions physiological factors governing parasite development in the thorax may be more important in limiting vectorial competence than the density of mf ingested.

Eosinophilia, anemia and parasitism in a rural region of northwest Thailand.

Parasitic infections are prevalent in Thailand. The parasites' relationship with anemia and eosinophilia has been rarely reported due to limited health care access, especially in rural areas. We studied the prevalence and relationship between eosinophilia, anemia and parasitism in 169 Thai-Karens from Mae Lamung and Mae Chan subdistricts, Umphang district, Tak Province, using an automated complete blood counter, and microscopic examination for intestinal parasites and microfilaria. In Mae Chan, 5 individuals were microfilaremic, 72% of individuals examined were infected with at least one kind of intestinal parasites, and 50% were anemic, with normal mean red cell volume (MCV). In Mae Lamung, 46% were parasitized (Triteeraprapab et al, 1997) but none were microfilaremic or anemic. In both populations, eosinophilia was prevalent (77%). Hookworm infection was found to be significantly associated with eosinophilia, but not anemia nor microcytosis of red cells.

Estudios comparativos clínicos y epidemiológicos de la oncocercosis entre dos grupos raciales del área hiperendémica de Ecuador / Comparative clinical and epidemiological studies between 2 racial groups in the hyperendemic area of onchocercosis in Ecuador

Se evaluó la importancia de los factores raciales en la epidemiología y las manifestaciones clínicas de la oncocercosis en la raza negra y la raza indígena Chachi, dos grupos étnicos diferentes del área hiperendémica del foco oncocercótico ecuatoriano. Los Chachi presentaron una significativa mayor densidad de microfilarias en la piel. De los negros, 72,2 por ciento presentaron lesiones oncocercóticas en la piel comparado con 56,4 por ciento en los Chachi. Se encontró mayor prevalencia de nódulos palpables en los Chachi (51,4 por ciento vs. 43,0 por ciento). La presencia de microfilarias en la cámara anterior del ojo fue más frecuente en los Chachi (51,9 por ciento vs. 34,2 por ciento), pero se evidenció una mayor prevalencia de corioretinitis y atrofia del nervio óptico en los negros (17,6 por ciento vs 11,2 por ciento y 8,5 por ciento vs. 0,5 por ciento, respectivamente)