Leishmania mexicana in a central Texas cat: clinical presentation, molecular identification, sandfly vector collection and novel management.

CASE SUMMARY: This case report documents the clinical appearance, diagnosis and novel treatment of a central Texas cat with cutaneous leishmaniosis. The cat presented with a linear erosion on the right pinnal margin, an ulcerated exophytic nodule on the right hock and a swelling in the right nostril. Cytological and histopathological findings were consistent with leishmaniosis. PCR confirmed the presence of Leishmania mexicana, a species endemic to Texas. An epidemiological investigation was conducted by trapping sandflies from the cat's environment. Sandflies collected were identified as Lutzomyia species, known vectors of Leishmania species. Given the lack of validated medical therapies for L mexicana in cats, treatments typically prescribed for canine leishmaniosis were administered. Allopurinol achieved clinical success but was discontinued due to suspected drug-related neutropenia. Topical imiquimod also improved lesional skin but was not sustainable due to application difficulty. Oral administration of artemisinin resulted in significant clinical improvement of cutaneous lesions without reported adverse events. Nearly 8 months after the initiation of artemisinin therapy, the cat remained systemically healthy with stable lesions. RELEVANCE AND NOVEL INFORMATION: This case report demonstrates endemic feline leishmaniosis in central Texas and provides the clinician with alternative therapeutic options for medical management.

Association of vectors and environmental conditions during the emergence of Peruvian horse sickness orbivirus and Yunnan orbivirus in northern Peru.

Since 1983, cases of diseased donkeys and horses with symptoms similar to those produced by alphaviruses were identified in two departments in northern Peru; however serological testing ruled out the presence of those viruses and attempts to isolate an agent were also unproductive. In 1997, also in northern Peru, two new orbiviruses were discovered, each recognized as a causative agent of neurological diseases in livestock and domestic animals and, at the same time, mosquitoes were found to be infected with these viruses. Peruvian horse sickness virus (PHSV) was isolated from pools of culicid mosquitoes, Aedes serratus and Psorophora ferox, and Yunnan virus (YUOV) was isolated from Aedes scapularis in the subtropical jungle (upper jungle) located on the slope between the east side of the Andes and the Amazonian basin in the Department of San Martín. Both viruses later were recovered from mosquitoes collected above the slope between the west side of the Andes and the coast (Department of Piura) in humid subtropical areas associated with the Piura River basin. In this region, PHSV was isolated from Anopheles albimanus and YUOV was isolated from Ae. scapularis. We discuss the ecology of vector mosquitoes during the outbreaks in the areas where these mosquitoes were found.

Molecular identification of sand flies (Diptera: Psychodidae) in eastern North America by using PCR-RFLP.

Sand flies (Diptera: Psychodidae) are small blood-feeding dipterans that are primary vectors of numerous human and livestock pathogens. Effective surveillance programs with accurate identification tools are critical in development and implementation of modern integrated pest management programs. Although morphological keys are available for North American species, identification can still be challenging owing to the nature of sample preparation and incompatibility with molecular or biochemical-based pathology assays. Further, the potential for introduction of Old World or other exotic species is not accounted for by current keys. Herein, we present the development and validation of a restriction fragment-length polymorphism-based molecular identification method. Specifically, cytochrome c oxidase subunit I, a mitochondrial DNA marker, was used to distinguish two species of adult sand flies indigenous to eastern North America with two exotic species not yet known to occur in the United States.

Hyperendemic malaria transmission in areas of occupation-related travel in the Peruvian Amazon.

BACKGROUND: Plasmodium vivax and Plasmodium falciparum cause a significant illness burden in Peru. Anopheline indices for populated communities in the peri-Iquitos region of Loreto have been reported to be remarkably low, with entomological inoculation rates (EIR) estimated at one to 30 infective bites per year based on a few studies in close proximity to the urban centre of Iquitos and surrounding deforested areas. Local reports suggest that a large number of the reported cases are contracted outside of populated communities in undeveloped riverine areas frequented by loggers and fishermen. METHODS: To better understand vectorial capacity in suspected high malaria transmission zones in a rural district near Iquitos, Peru, mosquito collections were conducted at different points in the seasonality of malaria transmission in 21 sites frequented by occupational labourers. Prevalence of Plasmodium spp in vectors was determined by circumsporozoite protein ELISA on individual mosquitoes. Slide surveillance was performed for humans encountered in the zone. RESULTS: In total, of 8,365 adult female mosquitoes examined, 98.5% were identified as Anopheles darlingi and 117 (1.4%) tested positive for sporozoites (P. falciparum, P. vivax VK210 or P. vivax VK247). Measured human biting rates at these sites ranged from 0.102 to 41.13 bites per person per hour, with EIR values as high as 5.3 infective bites per person per night. Six percent of the 284 blood films were positive for P. vivax or P. falciparum; however, 88% of the individuals found to be positive were asymptomatic at the time of sampling. CONCLUSIONS: The results of this study provide key missing indices of prominent spatial and temporal heterogeneity of vectorial capacity in the Amazon Basin of Peru. The identification of a target human subpopulation as a principal reservoir and dispersion source of Plasmodium species has important implications for vaccine development and the delivery of effective targeted malaria control strategies.

Detection of Rickettsia parkeri from within Piura, Peru, and the first reported presence of Candidatus Rickettsia andeanae in the tick Rhipicephalus sanguineus.

Domestic farm animals (n=145) were sampled for the presence of ectoparasites in northwestern Peru during March, 2008. Ninety domestic animals (62%) were positive for the presence of an ectoparasite(s) and produced a total collection of the following: 728 ticks [Amblyomma maculatum, Anocentor nitens, Rhipicephalus (Boophilus) microplus, Rhipicephalus sanguineus, and Otobius megnini], 12 lice (Haematopinus suis), and 3 fleas (Ctenocephalides felis). A Rickettsia genus-specific qPCR assay was performed on nucleic acid preparations of the collected ectoparasites that resulted in 5% (37/743, 35 ticks and 2 fleas) of the ectoparasites positive for the presence of Rickettsia. DNA from the positive individual ticks was tested with 2 other qPCR assays for the presence of the ompB gene in Candidatus Rickettsia andeanae or Rickettsia parkeri. Candidatus R. andeanae was found in 25 A. maculatum ticks and in two Rh. sanguineus ticks, whereas R. parkeri was detected in 6 A. maculatum ticks. Two A. maculatum were co-infected with both Candidatus R. andeanae and R. parkeri. Rickettsia felis was detected in 2 fleas, Ctenocephalides felis, by multilocus sequence typing of the 17-kD antigen and ompA genes. These findings expand the geographic range of R. parkeri to include Peru as well as expand the natural arthropod vector of Candidatus R. andeanae to include Rhipicephalus sanguineus.

Divergence of Lutzomyia (Psathyromyia) shannoni (Diptera: Psychodidae: Phlebotominae) is indicated by morphometric and molecular analyses when examined between taxa from the southeastern United States and southern Mexico.

The medically important sand fly Lutzomyia shannoni (Dyar 1929) was collected at eight different sites: seven within the southeastern United States and one in the state of Quintana Roo, Mexico. A canonical discriminant analysis was conducted on 40 female L. shannoni specimens from each of the eight collection sites (n = 320) using 49 morphological characters. Four L. shannoni specimens from each of the eight collection sites (n = 32) were sent to the Barcode of Life Data systems where a 654-base pair segment of the cytochrome c oxidase subunit 1 (CO1) genetic marker was sequenced from each sand fly. Phylogeny estimation based on the COI segments, in addition to genetic distance, divergence, and differentiation values were calculated. Results of both the morphometric and molecular analyses indicate that the species has undergone divergence when examined between the taxa of the United States and Quintana Roo, Mexico. Although purely speculative, the arid or semiarid expanse from southern Texas to Mexico City could be an allopatric barrier that has impeded migration and hence gene flow, resulting in different morphology and genetic makeup between the two purported populations. A high degree of intragroup variability was noted in the Quintana Roo sand flies.

Morphometric and molecular analyses of the sand fly species Lutzomyia shannoni (Diptera: Psychodidae: Phlebotominae) collected from seven different geographical areas in the southeastern United States.

A morphometric and molecular study of adult male and female Lutzomyia shannoni (Dyar 1929) collected at seven different locations within the southeastern United States was conducted to assess the degree of divergence between the grouped specimens from each location. The collection locations were as follows: Fort Bragg, NC; Fort Campbell, KY; Fort Rucker, AL; Ossabaw Island, GA; Patuxent National Wildlife Research Refuge, MD; Suwannee National Wildlife Refuge, FL; and Baton Rouge, LA. Forty males and forty females from each location were analyzed morphometrically from 54 and 49 character measurements, respectively. In addition, the molecular markers consisting of the partial cytochrome c oxidase subunit I (from 105 sand flies: 15 specimens/collection site) and the partial internal transcribed spacer 2 (from 42 sand flies: six specimens/collection site) were compared. Multivariate analyses indicate that the low degree of variation between the grouped specimens from each collection site prevents the separation of any collection site into an entity that could be interpreted as a distinct population. The molecular analyses were in concordance with the morphometric study as no collection location grouped into a separate population based on the two partial markers. The grouped specimens from each collection site appear to be within the normal variance of the species, indicating a single population in the southeast United States. It is recommended that additional character analyses of L. shannoni based on more molecular markers, behavioral, ecological, and physiological characteristics, be conducted before ruling out the possibility of populations or a cryptic species complex within the southeastern United States.

Population dynamics of Lutzomyia shannoni (Diptera: Psychodidae) at the Patuxent National Wildlife Research Refuge, Maryland.

The seasonal abundance and temporal patterns of the adult sand fly (Lutzomyia shannoni Dyar) were examined at the Patuxent National Wildlife Research Refuge, MD, from August 3, 2005, to July 29, 2006. A total of 138 (53 males, 85 females) L. shannoni was collected from 4 dry ice-baited traps set at the same 4 locations throughout the study. The male:female ratio was 1:2.4. All 4 traps, separated by a maximum distance of approximately 1.6 km, operated simultaneously on the collection dates. The collection dates were spaced apart by near weekly intervals during the months of expected sand fly activity. No collections occurred in December-February. August was clearly the period of peak adult abundance as the numbers collected were significantly greater during this month than any other month of collection. Results indicate the existence of a unimodal pattern of abundance with adult emergence beginning in June and ending by September. The temporal pattern and abundance differ from what has been observed for the species on Ossabaw Island, a barrier island located along coastal Georgia, and at San Felasco Hammock Preserve State Park, Gainesville, FL. Continued research is needed to compile multiyear data to confirm the temporal abundance patterns of this species in Maryland.

Morphological anomalies in two Lutzomyia (Psathyromyia) shannoni (Diptera: Psychodidae: Phlebotominae) specimens collected from Fort Rucker, Alabama, and Fort Campbell, Kentucky.

This report describes two male specimens of the sand fly species Lutzomyia shannoni (Dyar) (Diptera: Psychodidae: Phlebotominae) collected at Fort Rucker, AL, and Fort Campbell, KY, in dry ice-baited light traps during September 2005. The specimens were observed to have anomalies to the number of spines on the gonostyli. The taxonomic keys of Young and Perkins (Mosq. News 44: 263-285; 1984) use the number of spines on the gonostylus in the first couplet to differentiate two major groupings of North American sand flies. The two anomalous specimens were identified as L. shannoni based on the following criteria: (1) both specimens possess antennal ascoids with long, distinct proximal spurs (a near diagnostic character of L. shannoni in North America), (2) the sequences of the partial cytochrome c oxidase subunit 1 gene from both specimens indicated L. shannoni, and (3) the sequences of the internal transcribed spacer 2 molecular marker from both specimens indicated L. shannoni. The anomalous features are fundamentally different from each other as the Fort Rucker specimen possesses a fifth spine (basally located) on just one gonostylus, whereas the Fort Campbell specimen possesses five spines (extra spines subterminally located) on both gonostyli. Because the gonostyli are part of the external male genitalia, anomalies in the number of spines on the gonostyli may have serious biological consequences, such as reduced reproductive success, for the possessors. These anomalies are of taxonomic interest as the specimens could easily have been misidentified using available morphological keys.

Vigilancia de arbovirus en el departamento de Piura: Perú - 2008 / Surveillance of arbovirus in Piura department: Perú - 2008

La vigilancia virológica de vectores en el departamento de Piura se realiza considerando su variada topografía, amplia gama de climas en distancias cortas (desde desiertos hasta bosques tropicales) y la precipitación pluvial elevada durante la presentación de El Niño, favorecen el aumento de vectores de arbovirosis epizoodémicas, como la Encefalitis Equina Venezolana y los endémicos contra el Dengue. En el 2008 se realizó un muestro entomológico en tres distritos de la Pronvicia de Ayabaca (Paimas, Montero y Suyo). Este muestreo se viene desarrollando desde el año 1981, con la finalidad de determinar el desplazamiento de vectores de arbovirus occidentales. Durante la época del muestreo (estación lluviosa) se capturaron un total de 1916 ejemplares principalmente anofelinos. Las mayores capturas correspondieron a An. Albimanus y a Oc. Scapularis. No se recuperó ningún Arbovirus de los 50 pools de mosquitos inoculados en cultivos celulares C6/36 y Vero. Parece que las condiciones climáticas normales para la estación no constituyeron un riesgo para la presentación de vectores epizoóticos del VEE o del Bunyavirus asociados a epizootias durante los años 1972-73 y de 1983-84, respectivamente. No se encontró Ae. Aegypti, vector del dengue de amplia distribución en el departamento, probablemente porque las áreas de muestreo son rurales. Por otro lado, se observó la incursión del vector selvático de la fiebre amarilla Haemagogus soperi en la localidad de Sarayuyo, distrito de Suyo que hace necesario mantener la vigilancia entomológica.

Viral surveillance of vector-borne human diseases in the department of Piura is performed considering its varied topography, with a wide range of climates over short distances (from deserts to tropical forests) and the increase of rainfall mainly during the presentation of ENSO. This variability enables to study both the epizoodemic arbovirosis as Venezuelan Equine Encephalitis and those endemic as Dengue. In 2008, entomological sampling was performed looking arbovirus in three districts of the Province of Ayabaca (Paimas, Montero and Suyo). This sampling has been developing since 1981 in order to determine the displacement of arbovirus vectors in eastern tropical areas and western desert forests. During the sampling period (rainy season) were captured a total of 1916 specimens mainly anophelines. The largest catches were of An albimanus and Oc. Scapularis. Any arboviruses were not recovered from the 50 polls of mosquitos inoculated in C6/36 and Vero cell cultures. It seems that weather conditions were normal and did nor constituted a risk in order to increase epizootic vectors of VEE or Bunyavirus associated with epizootics in the years 1972-73 and 1983-84, respectively. Ae. Aegypti vector of Dengue widely distributed in the department, was not found probably because the sampled areas are rural. Furthermore, we observed the incursion of wild vector of yellow Fever. Haemogogus soperi, in the town of Sarayuyo, Suyo district, for this reason it is necessary to maintain entomological surveillance.

New state records for Lutzomyia shannoni and Lutzomyia vexator.

Two species of phlebotomine sand flies, Lutzomyia shannoni (Dyar) and Lutzomyia vexator (Coquillett), are reported for the first time from Kentucky and Ohio. L. vexator also is reported for the first time from Tennessee. These insects were found in a northeasterly band extending from southwestern Kentucky to southwestern Ohio. Both species were consistently captured from mid-July through September in 2006 and 2007 by using CO2-baited Center for Disease Control light traps. Weekly sampling revealed that these flies are more abundant in the southern part of this band than in the northern part, but increasing densities throughout this new range indicate that the flies are currently expanding their range. Although both species have been reported further north along the Atlantic coast, and L. vexator along the Pacific coast, neither of them had been reported this far north along the Mississippi Valley. Previous reports established L. shannoni as far north as west central Tennessee and L. vexator in a similar spatial pattern in the eastern part of its range, extending as far north as northern Alabama. Whether the new records reported herein represent a northerly expansion of the geographic range of these species or are reflective of sampling changes is inconclusive. However, the former scenario could presage an increased prevalence of the diseases associated with this group of insects.

Improved molecular technique for the differentiation of neotropical anopheline species.

We evaluated a PCR-RFLP of the ribosomal internal transcribed spacer 2 region (ITS2) to distinguish species of Anopheles commonly reported in the Amazon and validated this method using reared F1 offspring. The following species of Anopheles were used for molecular analysis: An. (Nys.) benarrochi, An. (Nys.) darlingi, An. (Nys.) nuneztovari, An. (Nys.) konderi, An. (Nys.) rangeli, and An. (Nys.) triannulatus sensu lato (s.l.). In addition, three species of the subgenus Anopheles, An. (Ano.) forattini, An. (Ano.) mattogrossensis, and An. (Ano.) peryassui were included for testing. Each of the nine species tested yielded diagnostic banding patterns. The PCR-RFLP method was successful in identifying all life stages including exuviae with small fractions of the sample. The assay is rapid and can be applied as an unbiased confirmatory method for identification of morphologic variants, disputed samples, imperfectly preserved specimens, and life stages from which taxonomic keys do not allow for definitive species determination.

Distribution of Ochlerotatus togoi along the Pacific coast of Washington.

Before this study, the mosquito Ochlerotatus (Finlaya) togoi (Theobald) had been reported from only 2 locations within the continental United States, both of which were documented in Washington State. This study used active and passive surveillance to determine the current distribution of Oc. togoi along the Pacific Coast of Washington. Results of the study show that small, but stable, populations of Oc. togoi exist in the northern San Juan Island region of Puget Sound. Geological formations in this region are conducive to rock holes and support populations of Oc. togoi. No members of Oc. togoi were found on the southwestern Washington coast, the coast of the Olympic Peninsula, or in the lower Puget Sound.