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A leishmaniose visceral (LV) é uma zoonose classificada como doença tropical negligenciada, atualmente não prevenível e transmitida por vetor artrópode. É um crescente problema global de saúde pública uma vez que as atuais medidas de controle baseadas, entre outras coisas, no diagnóstico e eliminação do reservatório canino, não têm sido capazes de reduzir a transmissão e a expansão territorial da doença. A presença de cães em área urbana, domiciliados ou comunitários, pode aumentar o risco para humanos, uma vez que cães são os principais reservatórios do parasita no ciclo urbano. A utilização de coleiras impregnadas com deltametrina a 4% em cães soronegativos, diagnosticados em inquéritos sorológicos caninos em área de transmissão, pode ser uma estratégia eficaz no controle da LV. Nesta pesquisa foi analisada a efetividade de uma estratégia alternativa de controle da doença fundamentada na substituição da eutanásia pela aplicação de coleira impregnada com deltametrina nos cães sororreagentes, identificados em inquéritos feitos em áreas de transmissão. A nova medida de controle, aqui denominada encoleiramento estratégico, foi empregada pelo Centro de Controle de Zoonoses de Bauru, SP entre os anos de 2018 e 2019. A avaliação da efetividade dessa estratégia foi feita por meio de uma análise prospectiva, comparando-se a incidência da presença de anticorpos anti-leishmania identificada pelos testes diagnósticos oficiais (TR-DPP e ELISA), entre os cães de dois bairros do município de Bauru, SP, sendo um deles onde se praticou a eutanásia de cães sororreagentes e outro somente, o aqui denominado, encoleiramento estratégico. Foram avaliados resultados de testes sorológicos aplicados em amostras de soro coletadas da totalidade dos cães dos dois bairros, em três tempos consecutivos, com intervalos de cerca de 200 dias entre eles. As comparações das incidências foram feitas entre as coortes, nos três tempos distintos, e dentro de cada coorte, nos mesmos tempos. A comparação entre as taxas de incidência de reatividade sorológica nos dois bairros durante o período do estudo mostrou que a medida de encoleiramento estratégico foi 4 vezes mais efetiva que a eutanásia na redução da soropositividade dos cães. Os resultados deste estudo mostram que o encoleiramento estratégico apresenta-se como uma medida promissora para ser empregada no controle da LV em cães de áreas endêmicas, em especial por prescindir de eutanásia dos animais infectados, um procedimento controverso e de impacto negativo na sociedade. Visceral leishmaniasis (VL) is a zoonosis classified as a neglected tropical disease, currently not preventable and transmitted by an arthropod vector. VL is a growing global public health problem because current control measures, based on diagnosis and elimination of the source of infection, among other things, have not been able to reduce the transmission and territorial expansion of the disease. The presence of domiciled, semi-domiciled or stray dogs in urban areas can increase the risk for humans, because dogs are the main reservoir of the parasite in the urban cycle. Recent studies have shown that the use of 4% deltamethrin-impregnated collars in seronegative dogs diagnosed in canine serological surveys carried out in the transmission area can be an effective strategy in the control of VL, however, in these studies the controversial euthanasia of positive dogs was part of the measures of control evaluated. In this study, an alternative strategy to control the disease was analyzed, based on replacing euthanasia with the application of a collar impregnated with deltamethrin in seroreactive dogs identified in surveys carried out in areas of transmission. The new strategy, here called strategical collaring, was employed by the Center for Zoonosis Control (CZC) in the municipality of Bauru, SP throughout 2018 and 2019. The evaluation of the effectiveness of this strategy was carried out through a prospective analysis, in order to compare the incidence of antibodies identified by official diagnostic tests (TR-DPP and ELISA) among dogs from two districts of the city of Bauru SP, one of them where the euthanasia of infected dogs was practiced as a method of VL control and the other, the strategical collaring. Results of serological tests applied to serum samples collected from all the dogs in the two neighborhoods were evaluated, in three consecutive times, with intervals of about 200 days between them. Comparisons of incidences were made between cohorts at three different times and within each cohort at the same times. The comparison between the incidence rates of serological reactivity in the two neighborhoods during the study period showed that the strategical collaring measure was up to 4 times more effective than euthanasia in reducing the seropositivity of dogs. The results of this study show that reverse collaring is a promising measure to be used in the control of VL in dogs in endemic areas, especially as it does not require euthanasia of infected animals, a controversial procedure with a negative impact on society.

Previous studies demonstrated that insecticide collars are highly effective in reducing canine visceral leishmaniasis (CVL); however, it is unclear if the efficacy differs by socioeconomic conditions across diverse communities. This study aimed fourfold: (i) to evaluate the protection of 4% impregnated deltamethrin collared (DMC) dogs in different areas of an endemic city for visceral leishmaniasis (VL); (ii) to analyze socioeconomic variables with the seroconversion rates; (iii) to analyze the indirect effect of DMC on untreated dogs in areas of intervention; and, (iv) to evaluate the potential transmission to other dogs in the same household when one positive dog is present. The study employed the municipality of Bauru, São Paulo, Brazil, as the area of interest and used Geographic Information System tools to fit binary logistic regression models. Dogs were divided into three cohort studies: intervention with DMC (I), indirect effect of DMC (IE), and control (C). Pre-intervention, lower mean income was associated with higher rates of CVL and a 142% increase in the odds of transmission (OR = 1.42, p-value = 0.001, CI 1.14,1.77). Post-intervention, lower-income areas depicted greater efficacy (76%) than higher-income areas (45%). The overall efficacy of DMC in preventing CVL was 63%; however, seroconversion rates were higher for IE (6.02%) than C (3.78%), revealing the failure of the indirect protection of DMC to manage the spread of the disease among the general non-wearing DMC canine population living in the same area. The protected dogs may repel the vectors, and non-protected dogs attract them, creating a higher transmission rate for non-protected dogs. Greater seroconversion was observed for living with an infected dog (10.20% in IE and 8.75% in C) than for the indirect effect of DMC, demonstrating the social burden of CVL. Overall, uncollared dogs have three times higher odds of being infected with CVL than DMC dogs (p 0.005), and uncollared dogs living with (an) infected dog(s) in the same household can reach 3.5 times higher odds than those living with negative ones (p 0.005). The results may assist in enhancing public policies and minimizing inequality in low and middle-income countries that suffer from neglected diseases such as VL.

BACKGROUND Dogs are the main peridomiciliary reservoir of Leishmania infantum thus the correct diagnosis of infection is essential for the control of the transmission and treatment as well. However, the diagnosis is based on serological assays that are not fully effective. OBJECTIVE We aimed to establish an effective serological assay for the diagnosis of L. infantum infected dogs using Leishmania-derived recombinant antigens. METHODS Leishmania derived rK39-, rK28-, rKR95-based enzyme-linked immunosorbent assay (ELISA) was standardized using symptomatic and asymptomatic L. infantum-infected dogs. Then 2,530 samples from inquiry in endemic areas for VL were evaluated and the results compared with recommended assays by the Brazilian Ministry of Health (MH algorithm). Further samples from a cohort of 30 dogs were searched. FINDINGS For rK39-, rK28- and rKR95-ELISA the sensitivity was around 97% and specificity 100%. The positivity of these three ELISA in the inquiry samples was 27-28%, around 10% higher than the assays currently in use. When cohort samples were searched, we observed likely false-negative results (> 65%) with supposedly negative samples that turned positive six months later with the assays in use (MH algorithm). MAIN CONCLUSIONS For the diagnosis of L. infantum-infected dogs, rK39-based ELISA showed better diagnostic performance than other assays in use in Brazil and worldwide.

Made available in DSpace on 2022-04-29T08:32:40Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-08-01 When it comes to visceral leishmaniasis (VL) in Brazil, one of the main targets of public health policies of surveillance is the control of domestic canine reservoirs of Leishmania infantum. This paper aims to evaluate the effect of the dog population and household environment for the maintenance of natural foci in the transmission to human and animal hosts in an endemic city for VL, Bauru, in Brazil. We collected 6, 578 blood samples of dogs living in 3, 916 households from Nov.2019 to Mar.2020 and applied geospatial models to predict the disease risk based on the canine population. We used Kernel density estimation, cluster analysis, geostatistics, and Generalized Additive Models (GAM). To validate our models, we used cross-validation and created a receiver operating characteristic (ROC) curve. We found an overall canine VL (CVL) seroprevalence of 5.6% for the sampled dogs, while for the households, the positivity rate was 8.7%. Odds ratios (OR) for CVL increased progressively according to the number of canines for >2 dogs (OR 2.70); households that already had CVL in the past increased the chances for CVL currently (OR 2.73); and the cases of CVL increase the chances for human VL cases (OR 1.16). Our models were statistically significant and demonstrated a spatial association between canine and human disease cases, mainly in VL foci that remain endemic. Although the Kernel density ratio map had the best performance (AUC = 82), all the models showed high risk in the city's northwest area. Canine population dynamics must be considered in public policies, and geospatial methods may help target priority areas and planning VL surveillance in low and middle-income countries. Parasitology and Mycology Center Adolfo Lutz Institute (IAL) Sao Paulo Adolfo Lutz Institute Regional Laboratories Center II Bauru, Bauru Center for Zoonoses Control of Bauru Health Secretariat of Bauru Bioterium Nucleos Adolfo Lutz Institute (IAL) Sao Paulo Institute of Geography Federal University of Uberlándia, Uberlándia Department of Geography Sao Paulo State University Faculty of Sciences and Technology (FCT/UNESP) Presidente Prudente Department of Geography Sao Paulo State University Faculty of Sciences and Technology (FCT/UNESP) Presidente Prudente

Hammondia heydorni is a cyst forming coccidia closely related to other apicomplexans, such as Toxoplasma gondii, Neospora caninum and Hammondia hammondi with a two-host life cycle. Dogs and other canids as red foxes (Vulpes vulpes) and coyotes (Canis latrans) may serve as definitive hosts for H. heydorni. Sporulated oocysts are infective for cattle, sheep and goats, which may serve as intermediate hosts. Herein, we describe the ability of crab-eating fox (Cerdocyon thous), a wild carnivore that is commonly found from northern Argentina to northern South America, to serve as definitive host of H. heydorni. The whole masseter muscle and brain from two 2-year-old bovines were collected, minced and pooled together for the fox infection. The bovine pooled tissues were equally administered to four foxes, in two consecutive days. Two foxes shed subspherical unsporulated oocysts measuring 10–15 μm, after 8 and 9 days post-infection, respectively. One of the foxes eliminated oocysts for 5 days, while the other fox shed oocysts for 9 days. A DNA sample of oocysts detected at each day of oocyst elimination was tested by two PCRs, one of them carried out employing primers directed to the common toxoplasmatiid 18S and 5.8S ribosomal RNA coding genes (PCR-ITS1) and the other based on heat-shock protein 70 kDa coding gene (PCR-HSP70). These samples were also submitted to a N. caninum specific nested-PCR protocol based on a N. caninum specific gene (Nc5-nPCR). All of them were positive by PCR-ITS1 and PCR-HSP70 but negative by Nc5-nPCR. The PCR-ITS1 and PCR-HSP70 nucleotide sequences amplified from the oocysts shed by the foxes revealed 100% identity with homologous sequences of H. heydorni. In conclusion, it is clear that H. heydorni also uses the crab-eating fox as a definitive host. The crab-eating fox is usually reported to live in close contact with livestock in several regions of Brazil. Therefore, it is reasonable to infer that such carnivores may play an important role in the sylvatic and domestic cycles of H. heydorni infection.

A space-time analysis of American visceral leishmaniasis (AVL) in humans in the city of Bauru, São Paulo State, Brazil was carried out based on 239 cases diagnosed between June 2003 and October 2008. Spatial analysis of the disease showed that cases occurred especially in the city's urban areas. AVL annual incidence rates were calculated, demonstrating that the highest rate occurred in 2006 (19.55/100,000 inhabitants). This finding was confirmed by the time series analysis, which also showed a positive tendency over the period analyzed. The present study allows us to conclude that the disease was clustered in the Southwest side of the city in 2006, suggesting that this area may require special attention with regard to control and prevention measures.

Neospora caninum e Hammondia heydorni são coccídeos formadores de cistos proximamente relacionados com outros organismos heteroxenos pertencentes ao filo Apicomplexa, como Toxoplasma gondii e Hammondia hammondi. Para saber se o cachorro-do-mato (Cerdocyon thous), um canídeo silvestre comumente encontrado do nordeste da Argentina ao nordeste da América do Sul, pode ser hospedeiro definitivo de N. caninum, exemplares de cachorro-do-mato foram alimentados com tecidos de bovinos e bubalinos naturalmente infectados por N. caninum. As fezes dos cachorros-do-mato foram colhidas e examinadas diariamente para pesquisa de oocistos. No primeiro experimento, masseter e cérebro (peça inteira dos dois órgãos) colhidos de dois bovinos de aproximadamente dois anos de idade, foram picados, misturados e fornecidos às quatro raposas em dois dias consecutivos. Duas raposas eliminaram oocistos sub-esféricos não esporulados medindo 10-13µm no oitavo e nono dias pós-infecção, respectivamente. Uma das raposas eliminou oocistos por cinco e a outra nove dias. Amostras de DNA extraída dos de oocistos de cada dia de eliminação foram analisadas por PCR com primers grupo-específicos PCR-ITS-1 e espécie-especificos Nc5-NPCR. Todas as amostras foram positivas por PCR-HSP-70 e PCR-ITS-1, mas negativas por Nc5-NPCR. A sequência de nucleotídeos PCR-ITS-1 revelou 100% de identidade com seqüências homólogas de H. Heydorni. Concluímos que H. heydorni também utiliza o cachorro-do-mato (Cerdocyon thous) como hospedeiro definitivo. O cachorro-do-mato é usualmente reportado em contato com rebanhos em diversas regiões do Brasil. Então, é razoável inferir que estes carnívoros possam exercer um importante papel nos ciclos silvestre e doméstico da infecção por H. heydorni. Em um segundo experimento, sete exemplares de cachorro-do-mato (Cerdocyon thous) foram alimentados com tecidos de 3 búfalos (Bubalus bubalis) naturalmente infectados por N. caninum. As fezes das raposas foram analisadas 30 dias antes e trinta depois da infecção experimental e nenhuma das raposas eliminou oocistos N. caninum-like nas fezes. Estes resultados sugerem que o cachorro-do-mato (Cerdocyon thous) não é hospedeiro definitivo de N. caninum, entretanto estudos adicionais são necessários. Neospora caninum and Hammondia heydorni are two cyst forming coccidians closely related to other apicomplexans, such as Toxoplasma gondii and Hammondia hammondi with a two host life-cycle. To clarify whether crab-eating fox (Cerdocyon thous), a wild carnivore commonly found from northern Argentina to northern South America, can be final hosts of N. caninum, foxes were fed on tissues of cows and water buffaloes naturally infected with N. caninum. Faeces were examined from 30 days before to 30 days after infection. In experiment 1, the whole masseter muscle and brain from two-year old bovines are collected, minced and pooled together for the fox infection. The bovine pooled tissues were equally administered for four fox, in two consecutive days. Two foxes shed unsporulated oocysts subspherical in shape measuring 10-13µm, after eight and nine days post-infection, respectively. One of the foxes eliminated oocysts for five days, while the other fox shed oocysts for nine days. A DNA sample of oocysts detected at each day of oocyst elimination was tested by a PCR carried out employing primers directed to the common toxoplasmatiid 18S and 5.8S coding genes (PCR-ITS-1). These samples were also submitted to a N. caninum specific gene (Nc-5nPCR). All of them were positive by PCR-ITS-1 and negative by Nc-5nPCR. The PCR-ITS-1 nucleotide sequence revealed 100% identity with homologous sequences of H. heydorni. In conclusion, it is clear that H. heydorni also uses the crab-eating fox as a definitive host. The crab-eating fox is usually reported to live in close contact with livestock in several regions of Brazil. Therefore it is reasonable to infer that such carnivores may play an important role in the sylvatic and domestic cycles of H. heydorni infection. In experiment 2, seven fox consumed tissues from three N. caninum naturally infected water buffaloes. Feaces were examined from 30 days before to 30 days after infection and none fox shed N. caninum-like oocysts. These results suggest that the crab-eating fox cannot be definitive host of N. caninum; however additional studies will be necessary.