RESUMO
INTRODUCTION: We examined whether the United Kingdom (UK) or the United States (US) screening criteria are more appropriate for retinopathy of prematurity (ROP) screening in Hong Kong, in terms of sensitivity for detecting type 1 ROP and the number of infants requiring screening. METHODS: In this retrospective cohort study, we reviewed the medical records of all infants who underwent ROP screening from 2009 to 2018 at a tertiary hospital in Hong Kong. During this period, all infants born at gestational age (GA) ≤31 weeks and 6 days or birth weight (BW) <1501 g (ie, the UK screening criteria) underwent ROP screening. We determined the number of infants requiring screening and the number of type 1 ROP cases that would have been missed if the US screening criteria (GA ≤30 weeks & 0 days or BW ≤1500 g) had been used. RESULTS: Overall, 796 infants were screened using the UK screening criteria. If the US screening criteria had been used, the number of infants requiring screening would have decreased by 21.1%; all type 1 ROP cases would have been detected (38/38, 100% sensitivity). Of the 168 infants who would not have been screened using the US screening criteria, only four of them (2.4%) had developed ROP (all maximum stage 1 only). CONCLUSION: In our population, the use of the US screening criteria could reduce the number of infants screened without compromising sensitivity for the detection of type 1 ROP requiring treatment. We suggest narrowing the GA criterion for consistency with the US screening criteria during ROP screening in Hong Kong.
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Retinopatia da Prematuridade , Humanos , Recém-Nascido , Peso ao Nascer , Idade Gestacional , Hong Kong/epidemiologia , Triagem Neonatal , Retinopatia da Prematuridade/diagnóstico , Retinopatia da Prematuridade/epidemiologia , Retinopatia da Prematuridade/terapia , Estudos Retrospectivos , Fatores de Risco , Reino Unido/epidemiologia , Estados Unidos/epidemiologiaAssuntos
Neoplasias da Mama , Humanos , Feminino , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Inibidores da Aromatase/farmacologia , Inibidores da Aromatase/uso terapêutico , Receptores de Estrogênio/genética , Receptores Androgênicos/genética , Resistencia a Medicamentos Antineoplásicos/genéticaRESUMO
Transgenesis for Strongyloides and Parastrongyloides was accomplished in 2006 and is based on techniques derived for Caenorhabditis elegans over two decades earlier. Adaptation of these techniques has been possible because Strongyloides and related parasite genera carry out at least one generation of free-living development, with adult males and females residing in soil contaminated by feces from an infected host. Transgenesis in this group of parasites is accomplished by microinjecting DNA constructs into the syncytia of the distal gonads of free-living females. In Strongyloides stercoralis, plasmid-encoded transgenes are expressed in promoter-regulated fashion in the F1 generation following gene transfer but are silenced subsequently. Stable inheritance and expression of transgenes in S. stercoralis requires their integration into the genome, and stable lines have been derived from integrants created using the piggyBac transposon system. More direct investigations of gene function involving expression of mutant transgene constructs designed to alter intracellular trafficking and developmental regulation have shed light on the function of the insulin-regulated transcription factor Ss-DAF-16. Transgenesis in Strongyloides and Parastrongyloides opens the possibility of powerful new methods for genome editing and transcriptional manipulation in this group of parasites. Proof of principle for one of these, CRISPR/Cas9, is presented in this review.
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Marcação de Genes/métodos , Técnicas de Transferência de Genes , Genômica/métodos , Nematoides/genética , Animais , Instabilidade GenômicaRESUMO
AIMS: Despite a largely successful 'zero COVID' policy in 2020, the COVID-19 pandemic disrupted routine cancer services in the city of Hong Kong. The aims of this study were to examine the trends in cancer incidence before and during the COVID-19 pandemic and estimate missed cancer diagnoses. MATERIALS AND METHODS: We used population-based data from the Hong Kong Cancer Registry 1983-2020 to examine the trends of age- and sex-standardised cancer incidence before and during the COVID-19 pandemic. We applied: (i) the annual average percentage change (AAPC) calculated using the Joinpoint regression model and (ii) the autoregressive integrated moving average (ARIMA) model to forecast cancer incidence rates in 2020. Missed cancer diagnoses in 2020 were estimated by comparing forecasted incidence rates to reported rates. A subgroup analysis was conducted by sex, age and cancer site. RESULTS: The cancer incidence in Hong Kong declined by 4.4% from 2019 to 2020 (male 8.1%; female 1.1%) compared with the long-term AAPC of 0.5% from 2005 to 2019 (95% confidence interval 0.3, 0.7). The gap between the reported and forecasted incidence for 2020 ranged from 5.1 to 5.7% (male 8.5%, 9.8%; female 2.3%, 3.5%). We estimated 1525-1596 missed cancer diagnoses (ARIMA estimate -98, 3148; AAPC 514, 1729) in 2020. Most missed diagnoses were in males (ARIMA 1361 [327, 2394]; AAPC 1401 [1353, 1460]), with an estimated 479-557 missed cases of colorectal cancer (ARIMA 112, 837; AAPC 518, 597) and 256-352 missed cases of prostate cancer (AAPC 231, 280; ARIMA 110, 594). CONCLUSION: The incidence of new cancer diagnoses declined in 2020 contrary to the long-term increase over the previous decades. Significantly lower diagnoses than expected were observed in males, particularly for colorectal and prostate cancers. Fewer reported cancer cases indicate missed diagnoses and could lead to delayed treatment that could impact future health outcomes.
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COVID-19 , Neoplasias , Humanos , Masculino , Feminino , Hong Kong/epidemiologia , COVID-19/epidemiologia , Pandemias , Neoplasias/diagnóstico , Neoplasias/epidemiologia , Previsões , IncidênciaRESUMO
Recent advances in molecular genetics and in imaging mean that it is now increasingly feasible to image biological processes within helminth parasites and to visualize interactions between worms and their hosts. Moreover, other innovative imaging approaches that are not dependent on transgenic parasites have been applied to, and or developed for, the study of helminth parasites and have provided novel and important insights into the biology of these important pathogens.
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Nematoides/citologia , Infecções por Nematoides , Trematódeos/citologia , Infecções por Trematódeos , Animais , Humanos , Microscopia Eletrônica , Nematoides/fisiologia , Infecções por Nematoides/parasitologia , Trematódeos/fisiologia , Infecções por Trematódeos/parasitologiaRESUMO
Background: Catastrophic antiphospholipid syndrome (CAPS) is an autoimmune thrombogenic disorder of small and large vessels caused by autoantibodies against phospholipids and phospholipid-binding proteins. This severe form of antiphospholipid syndrome (APS) presents clinically with simultaneous life-threatening multiorgan thrombosis and the presence of two or more persistent antiphospholipid antibodies (APL) confirmed on testing 12 weeks apart. Case Presentation. We describe a case report of a 66-year-old woman with detected antinuclear antibodies (ANA) pretransplant diagnosed with CAPS following orthotopic liver transplant. The patient had acute respiratory failure; Doppler ultrasound and CT angiogram confirmed thrombosis in the hepatic artery, subsequent occlusion of the jump graft, and a splenic infarct. Hypercoagulability workup showed elevated levels of anticardiolipin IgG and beta-2-glycoprotein IgG/IgM and positive lupus anticoagulant, treated with steroids and anticoagulation. The patient was discharged after one month and was transitioned from heparin to life-long warfarin. Conclusion: Our patient provided a standard presentation of CAPS with abnormal pretransplant levels of antinuclear antibodies (ANA). Although there have been studies investigating the relationship between anticardiolipin antibodies and lupus anticoagulants and APS, the relationship between pretransplant positive ANA or antimitochondrial antibodies (AMA) and CAPS has yet to be explored. Further studies will be needed to determine the significance of these antibodies. We recommend preoperative APL testing for patients with positive ANA and AMA at preliver transplant presentation.
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Ease of experimental gene transfer into viral and prokaryotic pathogens has made transgenesis a powerful tool for investigating the interactions of these pathogens with the host immune system. Recent advances have made this approach feasible for more complex protozoan parasites. By contrast, the lack of a system for heritable transgenesis in parasitic nematodes has hampered progress toward understanding the development of nematode-specific cellular responses. Recently, however, significant strides towards such a system have been made in several parasitic nematodes, and the possible applications of these in immunological research should now be contemplated. In addition, methods for targeted cell ablation have been successfully adapted from Caenorhabditis elegans methodology and applied to studies of neurobiology and behaviour in Strongyloides stercoralis. Together, these new technical developments offer exciting new tools to interrogate multiple aspects of the host-parasite interaction following nematode infection.
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Técnicas de Transferência de Genes , Interações Hospedeiro-Parasita , Nematoides/imunologia , Nematoides/fisiologia , Neurônios/parasitologia , Animais , Nematoides/genéticaRESUMO
Knockout mice deficient in tissue plasminogen activator (tPA) are protected against hippocampal excitotoxicity. But it is unknown whether similar neuroprotection occurs after transient global cerebral ischemia, which is known to selectively affect the hippocampus. In this study, we tested the hypothesis that hippocampal cell death in tPA knockout mice would be reduced after transient global cerebral ischemia, and this neuroprotection would occur concomitantly with amelioration of both intra- and extracellular proteolytic cascades. Wild-type and tPA knockout mice were subjected to 20 min of transient bilateral occlusions of the common carotid arteries. Three days later, Nissl and terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling staining demonstrated that hippocampal cell death was significantly reduced in tPA knockout brains compared with wild-type brains. Caspase-3 and the two major brain gelatinases (matrix metalloproteinase (MMP)-9 and MMP-2) were assessed as representative measurements of intra- and extracellular proteolysis. Post-ischemic levels of caspase-3, MMP-9 and MMP-2 were similarly reduced in tPA knockouts compared with wild-type hippocampi. Taken together, these data suggest that endogenous tPA contributes to hippocampal injury after cerebral ischemia, and these pathophysiologic pathways may involve links to aberrant activation of caspases and MMPs.
Assuntos
Hipocampo/patologia , Ataque Isquêmico Transitório/genética , Ataque Isquêmico Transitório/patologia , Neurônios/patologia , Ativador de Plasminogênio Tecidual/deficiência , Animais , Morte Celular/fisiologia , Modelos Animais de Doenças , Proteína Glial Fibrilar Ácida/metabolismo , Hipocampo/fisiopatologia , Marcação In Situ das Extremidades Cortadas/métodos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosfopiruvato Hidratase/metabolismoRESUMO
Brain vasculature acts in synergism with neurons to maintain brain function. This neurovascular coupling, or trophic coupling between cerebral endothelium and neuron, is now well accepted as a marker for mapping brain activity. Neurovascular coupling is most active in the perivascular region, in which there are ample opportunities for cell-cell interactions within the neurovascular unit. This trophic coupling between cells maintains neurovascular function and cellular plasticity. Recent studies have revealed that even adult brains contain multiple stem cells of various lineages, which may provide cellular plasticity through the process of differentiation among these stem cell populations. In this chapter, we provide an overview of the process by which neurovascular components contribute to cellular plasticity in the cerebral perivascular regions, focusing on mechanisms of cell-cell interaction in adult brain.
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Plasticidade Celular/fisiologia , Córtex Cerebral/citologia , Córtex Cerebral/fisiologia , Circulação Cerebrovascular/fisiologia , Neurônios/fisiologia , Animais , Comunicação Celular , Humanos , Microglia/fisiologiaRESUMO
Coarse structural nested mean models are tools to estimate treatment effects from longitudinal observational data with time-dependent confounding. There is, however, no guidance on how to specify the treatment effect model, and model misspecification can lead to bias. We derive a goodness-of-fit test based on modified overidentification restrictions tests for evaluating a treatment effect model, and show that our test statistic is doubly-robust in the sense that, with a correct treatment effect model, the test has the correct type-I error if either the treatment initiation model or a nuisance regression outcome model is correctly specified. In a simulation study we show that the test has correct type-I error and can detect model misspecification. We use the test to study how the timing of antiretroviral treatment initiation after HIV infection predicts the effect of one year of treatment in HIV-positive patients with acute and early infection.
RESUMO
There is increasing evidence that modulation of tumor hypoxia may improve therapy outcome. However, most preclinical data are derived from subcutaneous rather than orthotopic tumor models. We investigated the effect of the hypoxia-modulating agents nicotinamide and carbogen on tumor hypoxia, tumor blood perfusion, and proliferative activity in liver metastases of the murine colon carcinoma line C26a. In untreated C26a liver metastases, we observed a considerable amount of hypoxia, similar to the amount in liver metastases of patients with colorectal cancer. Compared to untreated mice, we observed a significantly smaller hypoxic fraction in the liver metastases of mice treated with nicotinamide and carbogen breathing as single treatments or in combination. In the group of mice that underwent carbogen breathing, perfusion was significantly lower than in the untreated group, but the decrease was only marginal. The proliferative activity was similar in all groups. In C26a subcutaneous tumors, a similar effect on hypoxia has been observed that was, however, combined with a decrease in proliferative activity. The different effects of nicotinamide and carbogen on parameters of the tumor microenvironment in liver metastases and subcutaneous tumors suggest that the host tissue influences the mechanism by which nicotinamide and carbogen exert their effects. Since tumor hypoxia may be a clinical problem in colorectal liver metastases, our results open possibilities for further research on the effect of hypoxia modifiers on colorectal liver metastases to improve therapy outcome.
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Dióxido de Carbono/administração & dosagem , Carcinoma/patologia , Carcinoma/secundário , Hipóxia Celular/efeitos dos fármacos , Neoplasias do Colo/patologia , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/secundário , Niacinamida/administração & dosagem , Oxigênio/administração & dosagem , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Combinação de Medicamentos , Feminino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Infection of cynomolgus monkeys with microfilariae (Mf) of Onchocerca lienalis was studied as a model for human ocular onchocerciasis. Normal monkeys and immunized monkeys were given intracorneal/subconjunctival, intracameral, or intravitreal injections of Mf or bovine serum albumin (control). Selected animals were given diethylcarbamazine citrate (DEC) orally (15 mg/kg) daily after the ocular infection. Following intravitreal challenge, living Mf and fibrinous exudates were visible by slit-lamp in both the anterior chamber and vitreous. Eosinophils and macrophages surrounded the Mf in the vitreous, with degranulated eosinophils adherent to the Mf. Eosinophils infiltrated the uvea and surrounded the retinal vessels. After intracorneal injection of Mf, living Mf were visible by slit-lamp biomicroscopy in the cornea for 3 days, with minimal inflammation of the corneas occurring over the 7 days after injection. Intracamerally injected Mf induced anterior uveitis. The extent of the inflammatory reactions was not substantially altered by DEC treatment following intraocular injection of Mf. In vitro proliferative responses of peripheral blood leukocytes to a crude Mf antigen were not observed in infected monkeys and proliferative responses to mitogen declined in these animals. Responses to mitogen were inhibited by addition of Mf antigen in vitro in normal monkeys. Circulating IgG antibodies were present in the sensitized, intracorneally, and intravitreally challenged animals. No obvious correlations were present between IgG antibody level and ocular inflammation.
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Oftalmopatias/patologia , Oncocercose/patologia , Animais , Câmara Anterior/parasitologia , Dietilcarbamazina/uso terapêutico , Modelos Animais de Doenças , Olho/patologia , Oftalmopatias/tratamento farmacológico , Oftalmopatias/imunologia , Macaca fascicularis , Microfilárias/isolamento & purificação , Onchocerca/isolamento & purificação , Oncocercose/tratamento farmacológico , Oncocercose/imunologia , Corpo Vítreo/parasitologiaRESUMO
Chorioretinitis due to onchocerciasis is a major cause of blindness, and the pathogenesis is poorly understood. We have developed an experimental model for onchocercal chorioretinitis using cynomolgus monkeys (Macaca fascicularis). Two normal monkeys and two monkeys which had received prior sensitization with subcutaneous injections of live Onchocerca lienalis microfilariae were given intravitreal injections of either 0, 10, 50 or 500 live microfilariae. Posterior segment changes included disc edema, venous engorgement, retinal vasculitis, intraretinal hemorrhage, and progressive retinal pigment epithelial (RPE) disturbances. Histopathological findings included perivascular infiltrates with eosinophils, eosinophilic choroiditis, and RPE hypertrophy, hyperplasia and loss of pigment. Microfilariae in the retina had no surrounding inflammation but were found adjacent to areas of RPE alterations. Overall the inflammatory reaction in the two unsensitized monkeys was more severe than that seen in the sensitized monkeys. The retinal appearance of the monkeys resembled that found in human onchocerciasis, and this model appears to be a promising one for future investigations.
Assuntos
Coriorretinite/patologia , Oncocercose/patologia , Animais , Coriorretinite/microbiologia , Corioide/patologia , Macaca fascicularis , Microfilárias/isolamento & purificação , Onchocerca/isolamento & purificação , Oncocercose/complicações , Retina/patologia , Corpo Vítreo/patologiaRESUMO
Hartley guinea pigs were injected with microfilariae (Mf) of Onchocerca lienalis as a model for acute inflammatory responses to Mf in human Onchocerca volvulus infection. IgG autoantibody reactive with a 3 M KCl extract of guinea pig cornea was detected by ELISA in the serum of guinea pigs injected with O. lienalis Mf three or more times sub-conjunctivally, or two or more times subcutaneously. Administration of the microfilaricides diethylcarbamazine citrate and ivermectin did not alter the proportion of animals expressing autoantibody or the mean autoantibody titer. The severity of acute corneal inflammatory reactions to Mf was similar in animals with and without circulating autoantibody. Although autoantibody responses did not correlate with acute corneal inflammatory reactions to dead Mf, the ability of Mf to induce formation of an antibody reactive with a component of autologous cornea suggests that autoimmune mechanisms might participate in chronic onchocercal lesions in the cornea, eg, sclerosing keratitis.
Assuntos
Anticorpos Anti-Helmínticos/imunologia , Autoanticorpos/imunologia , Doenças da Córnea/imunologia , Oncocercose/imunologia , Animais , Doenças da Córnea/tratamento farmacológico , Doenças da Córnea/parasitologia , Dietilcarbamazina/uso terapêutico , Ensaio de Imunoadsorção Enzimática , Cobaias , Imunoglobulina G/imunologia , Ivermectina/uso terapêutico , Ceratite/imunologia , Masculino , Microfilárias/imunologia , Onchocerca/imunologia , Oncocercose/tratamento farmacológicoRESUMO
Two G protein alpha subunit genes orthologous to gpa-2 and gpa-3 in Caenorhabditis elegans have been identified in the parasitic nematode, Strongyloides stercoralis. These genes mediate chemosensory signal transduction regulating dauer arrest in C. elegans. In the parasite, they represent candidate mediators for regulation of the choice between free-living and parasitic life cycles, the obligatory developmental arrest of infective larvae, and reactivation of development after infection. The (A+T) content of these genes is 72.2% for coding sequences, 90% for introns, and 84.1% for 5' and 3' flanking regions, requiring the use of low extension temperatures for long distance PCR. The possible significance of conserved structural motifs of these proteins is discussed.
Assuntos
Proteínas Heterotriméricas de Ligação ao GTP/genética , Strongyloides stercoralis/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , Sequência Conservada , DNA de Helmintos/química , DNA de Helmintos/genética , DNA de Helmintos/isolamento & purificação , Proteínas Heterotriméricas de Ligação ao GTP/química , Dados de Sequência Molecular , Filogenia , Reação em Cadeia da Polimerase , RNA de Helmintos/química , RNA de Helmintos/genética , RNA de Helmintos/isolamento & purificação , Técnica de Amplificação ao Acaso de DNA Polimórfico , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico , Strongyloides stercoralis/químicaRESUMO
An experimental model of human onchocercal keratitis was produced by injecting guinea pigs subconjunctivally with microfilariae (Mf) of Onchocerca lienalis. Actively motile O lienalis Mf spontaneously penetrated the central cornea and produced gray-white midstromal opacities, 0.2 to 0.4 mm in diameter, resembling those of human onchocercal punctate keratitis. Histologically, small foci of eosinophil and mononuclear cell infiltration associated with small pockets of interstitial edema were present in the central corneal stroma, with eosinophil infiltrates in the conjunctiva, episclera, limbus, and ciliary body. The severity of the punctate keratitis was increased by repeated subconjunctival inoculations of Mf. Punctate lesions were not seen following subconjunctival injection of Mf in animals previously hyperimmunized by three subcutaneous injections of Mf. Hyperimmunization may produce an immune response capable of destroying the subconjunctivally injected Mf and preventing their migration into the central cornea.
Assuntos
Ceratite/etiologia , Onchocerca/patogenicidade , Oncocercose/etiologia , Animais , Túnica Conjuntiva , Córnea/parasitologia , Córnea/patologia , Modelos Animais de Doenças , Feminino , Cobaias , Imunização , Ceratite/parasitologia , Ceratite/patologia , Microfilárias/patogenicidade , Oncocercose/parasitologia , Oncocercose/patologia , Fatores de TempoRESUMO
BALB/cBYJ mice were immunized against larval Onchocerca volvulus by subcutaneous injection of normal, irradiated, or freeze-thaw-killed Onchocerca sp. larvae. The mice received challenge infections of O. volvulus third-stage larva (L3) contained in diffusion chambers implanted subcutaneously. At two-weeks postinfection, the diffusion chambers were removed and larval survival was assessed. When mice were immunized a single time with 35-krad-irradiated or normal O. volvulus L3, there was a significant reduction in the survival of challenge parasites. However, there was little or no reduction in challenge worm survival when mice were immunized a single time with freeze-thaw-killed O. volvulus L3 or fourth-stage larva (L4), or irradiated O. lienalis L3. When a second dose of freeze-thaw killed O. volvulus L3 or irradiated O. lienalis L3 was administered, there was a significant reduction in parasite survival in immunized mice. Immunization with O. volvulus L4 or a combination of L3 and L4 failed to confer protection. These results demonstrate that mice can be immunized against larval O. volvulus and that diffusion chambers are an efficient method for studying protective immunity to this parasite in a mouse model.
Assuntos
Onchocerca volvulus/imunologia , Oncocercose/prevenção & controle , Animais , Cultura em Câmaras de Difusão , Modelos Animais de Doenças , Imunização , Imunização Secundária , Injeções Subcutâneas , Larva/imunologia , Larva/efeitos da radiação , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Onchocerca volvulus/efeitos da radiação , Oncocercose/imunologia , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/imunologiaRESUMO
The minimal intestinal dose of an enzootic strain of Venezuelan encephalitis (VE) virus for Culex (Melanoconion) taeniopus mosquitoes caught at a marsh habitat of VE virus in Guatemala was less than five plaque forming units (pfu) of virus. Ingestion of this dose of virus in blood of viremic hamsters resulted in transmission of virus to other hamsters. This low intestinal threshold of an enzootic strain of VE virus indicates that the natural Guatemalan population of Cu. (Mel.) taeniopus can acquire VE virus from vertebrates that have viremia levels as low as 1,000-5,000 pfu/ml of blood, provided other factors do not limit virus interchange between mosquitoes and vertebrates.
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Culex/microbiologia , Vírus da Encefalite Equina Venezuelana/patogenicidade , Encefalomielite Equina/transmissão , Encefalomielite Equina Venezuelana/transmissão , Animais , Cricetinae , Feminino , Intestinos/microbiologiaRESUMO
Of three species of Nearctic black flies tested, Simulium pictipes Hagen proved the most efficient laboratory vector of the bovine parasite Onchocerca lienalis. Among flies inoculated intrathoracially with 40 microfilariae, numbers of 3rd-stage larvae per fly were 7.63 for S. pictipes, 7.54 for S. vittatum Zetterstedt, and 0.83 for S. decorum Walker. S. pictipes survived the longest under laboratory conditions, with 83.3% of the females remaining alive 10 days after inoculation with 40 microfilariae of O. lienalis. Using an artificial membrane feeding system, S. pictipes could be routinely infected with O. lienalis by mouth. This black fly was also susceptible to infection with the Guatemalan strain of O. volvulus. Among flies injected with 10 microfilariae the rate of infection with 3rd-stage larvae was 93%, with a mean of three 3rd-stage larvae per fly. Successful techniques for the large-scale recovery and cryopreservation of 3rd-stage larvae of O. lienalis were also developed. A motility rate of 92.7% was observed in larvae cryopreserved within vector black flies.