Ancylostoma ceylanicum,a re-emerging but neglected parasitic zoonosis

Although Ancylostoma ceylanicum is known to be an endemic and widely distributed hookworm of dogs and cats in Asia, its contribution to human morbidity as a potentially zoonotic hookworm remains largely unexplored. Since its discovery by Lane (1913) as a ‘new parasite’ of humans a century ago, the hookworm has been regarded as a ‘rare’ and ‘abnormal’ parasite and largely overlooked in surveys of human parasites. Recent molecular-based surveys in Asia, however, have demonstrated that A. ceylanicum is the second most common hookworm species infecting humans, comprising between 6% and 23% of total patent hookworm infections. In experimentally induced infections, A. ceylanicum mimics the clinical picture produced by the anthroponotic hookworms of ‘ground itch’ and moderate to severe abdominal pain in the acute phase. Natural infections with A. ceylanicum in humans have been reported in almost all geographical areas in which the hookworm is known to be endemic in dogs and cats, however for the majority of reports, no clinical data are available. Much like the anthroponotic hookworm species, patent A. ceylanicum adults can isolate within the jejunum to produce chronic infections that on occasion, may occur in high enough burdens to produce anaemia. In addition, the hookworm can act much like Ancylostoma caninum and be found lower in the gastrointestinal tract leading to abdominal distension and pain, diarrhoea and occult blood in the faeces accompanied by peripheral eosinophilia. Whether A. ceylanicum is capable of producing both classical hookworm disease and evoking morbidity through an uncontrolled allergic response in some individuals remains unascertained. Future investigations combining the use of molecular diagnostic tools with clinical and pathological data will shed further light on its role as a human pathogen. The control of this zoonosis necessitates an integrated and inter-sectorial “One Health” approach be adopted in communities where large numbers of dogs share a close relationship with humans.     

Fecal and Molecular Survey of Neospora caninum in Farm and Household Dogs in Mashhad Area, Khorasan Province, Iran

Neospora caninum is an important cause of abortion in dairy cattle worldwide. Dog is the definitive host for N. caninum and can infect dairy cattle. The aim of this study is to determine the prevalence of Neospora oocysts in feces of dogs from dairy farms. A total of 174 fecal samples was collected from 89 farm dogs and 85 household dogs during 2006 and 2008. Fecal samples of dogs were microscopically examined for detecting Hammondia Neospora-like oocysts (HNLO) by Mini Parasep®SF fecal parasite concentrator. HNLO were microscopically detected in 4 fecal samples (2.2%). The fecal samples with HNLO were examined by N. caninum-specific PCR. Two of the samples were positive for N. caninum. The 2 positive fecal samples were selected for inoculation to calves. Two inoculated calves were seronegative by ELISA for 4 months post-infection. This is the first report of finding N. caninum DNA in feces of farm dogs in Mashhad area, Iran.     

Detection of Helminth Eggs and Identification of Hookworm Species in Stray Cats,Dogs and Soil from Klang Valley,Malaysia

The present study was conducted to determine the prevalence of helminth eggs excreted in the faeces of stray cats, dogs and in soil samples. A total of 505 fresh samples of faeces (from 227 dogs and 152 cats) and soil were collected. The egg stage was detected via microscopy after the application of formalin–ether concentration technique. Genomic DNA was extracted from the samples containing hookworm eggs and used for further identification to the species level using real-time polymerase chain reaction coupled with high resolution melting analysis. Microscopic observation showed that the overall prevalence of helminth eggs among stray cats and dogs was 75.7% (95% CI = 71.2%–79.9%), in which 87.7% of dogs and 57.9% of cats were infected with at least one parasite genus. Five genera of heliminth eggs were detected in the faecal samples, including hookworms (46.4%), Toxocara (11.1%), Trichuris (8.4%), Spirometra (7.4%) and Ascaris (2.4%). The prevalence of helminth infections among stray dogs was significantly higher than that among stray cats (p < 0.001). Only three genera of helminths were detected in soil samples with the prevalence of 23% (95% CI = 15.1%–31%), consisting of hookworms (16.6%), Ascaris (4%) and Toxocara (2.4%). The molecular identification of hookworm species revealed that Ancylostoma ceylanicum was dominant in both faecal and soil samples. The dog hookworm, Ancylostoma caninum, was also detected among cats, which is the first such occurrence reported in Malaysia till date. This finding indicated that there was a cross-infection of A. caninum between stray cats and dogs because of their coexistent within human communities. Taken together, these data suggest the potential role of stray cats and dogs as being the main sources of environmental contamination as well as for human infections.     

Leishmania infantum xenodiagnosis from vertically infected dogs reveals significant skin tropism

BackgroundDogs are the primary reservoir for human visceral leishmaniasis due to Leishmania infantum. Phlebotomine sand flies maintain zoonotic transmission of parasites between dogs and humans. A subset of dogs is infected transplacentally during gestation, but at what stage of the clinical spectrum vertically infected dogs contribute to the infected sand fly pool is unknown.Methodology/Principal findingsWe examined infectiousness of dogs vertically infected with L. infantum from multiple clinical states to the vector Lutzomyia longipalpis using xenodiagnosis and found that vertically infected dogs were infectious to sand flies at differing rates. Dogs with mild to moderate disease showed significantly higher transmission to the vector than dogs with subclinical or severe disease. We documented a substantial parasite burden in the skin of vertically infected dogs by RT-qPCR, despite these dogs not having received intradermal parasites via sand flies. There was a highly significant correlation between skin parasite burden at the feeding site and sand fly parasite uptake. This suggests dogs with high skin parasite burden contribute the most to the infected sand fly pool. Although skin parasite load and parasitemia correlated with one another, the average parasite number detected in skin was significantly higher compared to blood in matched subjects. Thus, dermal resident parasites were infectious to sand flies from dogs without detectable parasitemia.Conclusions/SignificanceTogether, our data implicate skin parasite burden and earlier clinical status as stronger indicators of outward transmission potential than blood parasite burden. Our studies of a population of dogs without vector transmission highlights the need to consider canine vertical transmission in surveillance and prevention strategies.     

Comparative Experimental Infection Study in Dogs with Ehrlichia canis,E. chaffeensis,Anaplasma platys and A. phagocytophilum

Dogs acquire infections with the Anaplasmataceae family pathogens, E. canis, E. chaffeensis, E. ewingii, A. platys and A. phagocytophilum mostly during summer months when ticks are actively feeding on animals. These pathogens are also identified as causing diseases in people. Despite the long history of tick-borne diseases in dogs, much remains to be defined pertaining to the clinical and pathological outcomes of infections with these pathogens. In the current study, we performed experimental infections in dogs with E. canis, E. chaffeensis, A. platys and A. phagocytophilum. Animals were monitored for 42 days to evaluate infection-specific clinical, hematological and pathological differences. All four pathogens caused systemic persistent infections detectible throughout the 6 weeks of infection assessment. Fever was frequently detected in animals infected with E. canis, E. chaffeensis, and A. platys, but not in dogs infected with A. phagocytophilum. Hematological differences were evident in all four infected groups, although significant overlap existed between the groups. A marked reduction in packed cell volume that correlated with reduced erythrocytes and hemoglobin was observed only in E. canis infected animals. A decline in platelet numbers was common with E. canis, A. platys and A. phagocytophilum infections. Histopathological lesions in lung, liver and spleen were observed in all four groups of infected dogs; infection with E. canis had the highest pathological scores, followed by E. chaffeensis, then A. platys and A. phagocytophilum. All four pathogens induced IgG responses starting on day 7 post infection, which was predominantly comprised of IgG2 subclass antibodies. This is the first detailed investigation comparing the infection progression and host responses in dogs after inoculation with four pathogens belonging to the Anaplasmataceae family. The study revealed a significant overlap in clinical, hematological and pathological changes resulting from the infections.     

Genotyping of Enterocytozoon bieneusi in Farmed Blue Foxes (Alopex lagopus) and Raccoon Dogs (Nyctereutes procyonoides) in China

Enterocytozoon bieneusi is the most common species of microsporidia found both in humans and animals. Farmed animals, particularly closely associated to humans, may play an important role of zoonotic reservoir in transmitting this disease to humans. The fur industry is a major economic component in some parts of China. To understand the prevalence, genotype variety and zoonotic risk of E. bieneusi in farmed foxes and raccoon dogs, two species of fur animals, fecal specimens of 110 blue foxes and 49 raccoon dogs from Heilongjiang and Jilin provinces in China were examined by internal transcribed spacer (ITS)-based PCR. E. bieneusi was detected in 16.4% (18/110) blue foxes and 4.1% (2/49) raccoon dogs. Altogether, four genotypes of E. bieneusi were identified, including two known genotypes D (n = 13) and EbpC (n = 5), and two novel genotypes named as CHN-F1 (n = 1) in a fox and CHN-R1 (n = 1) in a raccoon dog. Phylogenetic analysis revealed that all the four genotypes were the members of zoonotic group 1. Genotypes D and EbpC were found in humans previously. The findings of zoonotic genotypes of E. bieneusi in the foxes and raccoon dogs suggest these animals infected with E. bieneusi may pose a threat to human health.     

Seroprevalence and associated risk factors of Toxoplasma gondii and Neospora caninum infections in cattle in Central India

Toxoplasma gondii and Neospora caninum are closely related cyst-forming parasites identified as important causes of reproductive failures in ruminants. While these parasites have been reported worldwide, seroprevalence and associated risk factors for cattle infections have not been determined in India. A total of 576 serum samples of cattle were analyzed for antibodies to T. gondii and N. caninum using enzyme-linked immunosorbent assay (ELISA), modified/Neospora agglutination test (MAT/NAT), and an indirect fluorescent antibody test (IFAT-tachyzoite and bradyzoite). Additionally, general information about cattle, movement of cats and dogs, the menace of rodents, management, and reproductive disorders were assessed to identify the potential risk factors. Overall, 32.9% (190/576) serum samples reacted positively to T. gondii and 24.8% (143/576) to N. caninum. The performance of the diagnostic tests showed excellent agreement between IFAT and ELISA (kappa [κ] = 0.98) and between MAT/NAT and ELISA (κ = 0.97). Combining both infections on avidity test, 94% sera had high-IgG avidity, and 3% had low-IgG avidity antibodies, indicating chronic infection in the majority of the cases. The identified risk factors (p < 0.05) for exposure to T. gondii were: increasing age (Odds Ratio [OR]: 2.02), movement of cat (OR: 4.8) and rodents (OR: 1.57) in the farm; and for N. caninum: increasing age (OR: 1.6), movement of dogs in the farm (OR: 2.07), drinking pond water (OR: 1.64) and abortion (OR: 1.82). These findings revealed that T. gondii and N. caninum infections are widespread in the study area and suggest conducting nationwide epidemiological studies owing to their economic importance.     

Detection of Neospora caninum DNA by polymerase chain reaction in bats from Southern China

Neospora caninum is an intracellular protozoan that infects many domestic and wild animals. Domestic dogs and other canids function as definitive hosts, while other mammals serve as natural intermediate hosts. In the present study, the brain tissues of bats collected in Yunnan Province, Southern China were tested by N. caninum specific-nested PCR, targeting the Nc-5 gene and the internal transcribed spacer 1 (ITS1) region of the ribosomal DNA to determine whether bats could be infected with N. caninum. N. caninum DNA was detected in 1.8% (4/227) of bats, i.e., 1.7% (1/60) in Rousettus leschenaultia, 1.7% (1/58) in Hipposideros pomona, 2.9% (2/69) in Rhinolophus pusillus, and none (0/40) in Myotis daubentoniid. The findings of the present study are only the first indication that bats could serve as an intermediate host, and further studies are necessary to confirm whether bats are involved in the transmission of N. caninum infections.     

Enhanced attraction of sand fly vectors of Leishmania infantum to dogs infected with zoonotic visceral leishmaniasis

BackgroundThe sand fly Phlebotomus perniciosus is the main vector of Leishmania infantum, etiological agent of zoonotic visceral leishmaniasis in the Western Mediterranean basin. Dogs are the main reservoir host of this disease. The main objective of this study was to determine, under both laboratory and field conditions, if dogs infected with L. infantum, were more attractive to female P. perniciosus than uninfected dogs.Methodology/Principal findingsWe carried out a series of host choice experiments and found that infected dogs were significantly more attractive to P. perniciosus than uninfected dogs in the laboratory as well as in the field. Significantly more P. perniciosus fed on infected dogs than on uninfected dogs. However, the fecundity of P. perniciosus fed on infected dogs was adversely impacted compared to uninfected dogs by lowering the number of laid eggs. Phlebotomus perfiliewi, the second most abundant sand fly species in the field site and a competent vector of L. infantum had similar trends of attractivity as P. perniciosus toward infected dogs under field conditions.ConclusionsThe results strongly suggest that L. infantum causes physiological changes in the reservoir host which lead to the host becoming more attractive to both male and female P. perniciosus. These changes are likely to improve the chance of successful transmission because of increased contact with infected hosts and therefore, infected dogs should be particularly targeted in the control of zoonotic visceral leishmaniasis in North Africa.     

Transcriptional changes in the hookworm, Ancylostoma caninum, during the transition from a free-living to a parasitic larva

Background

Third-stage larvae (L3) of the canine hookworm, Ancylostoma caninum, undergo arrested development preceding transmission to a host. Many of the mRNAs up-regulated at this stage are likely to encode proteins that facilitate the transition from a free-living to a parasitic larva. The initial phase of mammalian host invasion by A. caninum L3 (herein termed “activation”) can be mimicked in vitro by culturing L3 in serum-containing medium.

Methodology/Principal Findings

The mRNAs differentially transcribed between activated and non-activated L3 were identified by suppression subtractive hybridisation (SSH). The analysis of these mRNAs on a custom oligonucleotide microarray printed with the SSH expressed sequence tags (ESTs) and publicly available A. caninum ESTs (non-subtracted) yielded 602 differentially expressed mRNAs, of which the most highly represented sequences encoded members of the pathogenesis-related protein (PRP) superfamily and proteases. Comparison of these A. caninum mRNAs with those of Caenorhabditis elegans larvae exiting from developmental (dauer) arrest demonstrated unexpectedly large differences in gene ontology profiles. C. elegans dauer exiting L3 up-regulated expression of mostly intracellular molecules involved in growth and development. Such mRNAs are virtually absent from activated hookworm larvae, and instead are over-represented by mRNAs encoding extracellular proteins with putative roles in host-parasite interactions.

Conclusions/Significance

Although this should not invalidate C. elegans dauer exit as a model for hookworm activation, it highlights the limitations of this free-living nematode as a model organism for the transition of nematode larvae from a free-living to a parasitic state.     

Canine leishmaniosis in Tunisia: Growing prevalence,larger zones of infection

BackgroundDiscovered by Nicolle and Comte in 1908 in Tunisia, Leishmania infantum is an intracellular protozoan responsible for zoonotic canine leishmaniosis (CanL) and zoonotic human visceral leishmaniasis (HVL). It is endemic in several regions of the world, including Tunisia, with dogs considered as the main domestic reservoir. The geographic expansion of canine leishmaniosis (CanL) has been linked to global environmental changes that have affected the density and the distribution of its sand fly vectors.Methodology/Principal findingsIn this study, a cross-sectional epidemiological survey on CanL was carried out in 8 localities in 8 bioclimatic areas of Tunisia. Blood samples were taken from 317 dogs after clinical examination. Collected sera were tested by indirect fluorescent antibody test (IFAT; 1:80) for the presence of anti-Leishmania infantum antibodies. The overall seroprevalence was 58.3% (185/317). Among positive dogs, only 16.7% showed clinical signs suggestive of leishmaniosis. Seroprevalence rates varied from 6.8% to 84.6% and from 28% to 66% by bioclimatic zone and age group, respectively. Serological positivity was not statistically associated with gender. The presence of Leishmania DNA in blood, using PCR, revealed 21.2% (64/302) prevalence in dogs, which varied by bioclimatic zone (7.3% to 31%) and age group (7% to 25%). The entomological survey carried out in the studied localities showed 16 species of the two genera (Phlebotomus and Sergentomyia). P. perniciosus, P. papatasi, and P. perfiliewi were the most dominant species with relative abundances of 34.7%, 25% and 20.4%, respectively.Conclusions/SignificanceThe present report suggests a significant increase of CanL in all bioclimatic areas in Tunisia and confirms the ongoing spread of the infection of dogs to the country’s arid zone. Such an expansion of infection in dog population could be attributed to ecological, agronomic, social and climatic factors that affect the presence and density of the phlebotomine vectors.     

Tick-borne relapsing fever Borreliosis,a major public health problem overlooked in Senegal

BackgroundTick-borne relapsing fever (TBRF) is the most common vector-borne bacterial disease in humans in West Africa. It is frequently clinically confused with malaria. Our study aims to determine, on a micro-geographic scale, the conditions for the maintenance and spread of TBRF in the Niakhar district of Senegal.Methodology/Principal findingsWe conducted clinical, entomological and animal reservoir investigations. Field surveys were carried out in order to investigate the presence of Ornithodoros sonrai vector ticks and to detect Borrelia spp. by qPCR using the 16S rRNA and glpQ genes, respectively. Micromammal trapping series were carried out inside homes and Borrelia infection was detected using brain tissue qPCR. Capillary blood samples from febrile patients were also tested for Borrelia using qPCR. More than 97% (40/41) of the villages surveyed were infested with O. sonrai ticks. The prevalence of Borrelia spp. infections in ticks was 13% (116/910), and over 73% (85/116) were positively confirmed as being Borrelia crocidurae. Borreliosis cases accounted for 12% (94/800) of episodes of fever and all age groups were infected, with children and young people between the ages of 8–14 and 22–28 being the most infected by the disease (16% and 18.4%). TBRF cases occurred in all seasons, with a peak in August. In two species of small rodents that were found to be infected (Arvicanthis niloticus, Mus musculus), the proportion of Borrelia infection was 17.5% (10/57), and the highest prevalence of infection (40.9%, 9/22) was observed in A. niloticus.Conclusion/SignificanceOur study indicates that TBRF is an endemic disease in the Niakhar district, where children and young people are the most infected. Arvicanthis niloticus and O. sonrai ticks are massively present and appear to be the main epidemiological reservoirs causing its extensive spread to humans.     

Long-term impact of a ten-year intervention program on human and canine Trypanosoma cruzi infection in the Argentine Chaco

BackgroundInterruption of domestic vector-borne transmission of Trypanosoma cruzi is still an unmet goal in several American countries. In 2007 we launched a long-term intervention program aimed to suppress house infestation with the main domestic vector in southern South America (Triatoma infestans) and domestic transmission in Pampa del Indio, a resource-constrained, hyperendemic municipality with 1446 rural houses inhabited by Creole and indigenous people, in the Argentine Chaco ecoregion. Here, we assessed whether the 10-year insecticide-based program combined with community mobilization blocked vector-borne domestic transmission of T. cruzi to humans and dogs.MethodsWe carried out two municipality-wide, cross-sectional serosurveys of humans and dogs (considered sentinel animals) during 2016–2017 to compare with baseline data. We used a risk-stratified random sampling design to select 273 study houses; 410 people from 180 households and 492 dogs from 151 houses were examined for antibodies to T. cruzi using at least two serological methods.ResultsThe seroprevalence of T. cruzi in children aged <16 years was 2.5% in 2017 (i.e., 4- to 11-fold lower than before interventions). The mean annual force of child infection (λ) sharply decreased from 2.18 to 0.34 per 100 person-years in 2017. One of 102 children born after interventions was seropositive for T. cruzi; he had lifetime residence in an apparently uninfested house, no outside travel history, and his mother was T. cruzi-seropositive. No incident case was detected among 114 seronegative people of all ages re-examined serologically. Dog seroprevalence was 3.05%. Among native dogs, λ in 2016 (1.21 per 100 dog-years) was 5 times lower than at program onset. Six native adult dogs born after interventions and with stable lifetime residence were T. cruzi-seropositive: three had exposure to T. infestans at their houses and one was an incident case.ConclusionsThese results support the interruption of vector-borne transmission of T. cruzi to humans in rural Pampa del Indio. Congenital transmission was the most likely source of the only seropositive child born after interventions. Residual transmission to dogs was likely related to transient infestations and other transmission routes. Sustained vector control supplemented with human chemotherapy can lead to a substantial reduction of Chagas disease transmission in the Argentine Chaco.     

Flavonoids Modulate the Proliferation of Neospora caninum in Glial Cell Primary Cultures

Neospora caninum (Apicomplexa; Sarcocystidae) is a protozoan that causes abortion in cattle, horses, sheep, and dogs as well as neurological and dermatological diseases in dogs. In the central nervous system of dogs infected with N. caninum, cysts were detected that exhibited gliosis and meningitis. Flavonoids are polyphenolic compounds that exhibit antibacterial, antiparasitic, antifungal, and antiviral properties. In this study, we investigated the effects of flavonoids in a well-established in vitro model of N. caninum infection in glial cell cultures. Glial cells were treated individually with 10 different flavonoids, and a subset of cultures was also infected with the NC-1 strain of N. caninum. All of the flavonoids tested induced an increase in the metabolism of glial cells and many of them increased nitrite levels in cultures infected with NC-1 compared to controls and uninfected cultures. Among the flavonoids tested, 3'',4''-dihydroxyflavone, 3'',4'',5,7-tetrahydroxyflavone (luteolin), and 3,3'',4'',5,6-pentahydroxyflavone (quercetin), also inhibited parasitophorous vacuole formation. Taken together, our findings show that flavonoids modulate glial cell responses, increase NO secretion, and interfere with N. caninum infection and proliferation.     

Seroprevalence of Cryptosporidium parvum and Neospora caninum in cattle in the southern Kyushu region of Japan

Cryptosporidium parvum and Neospora caninum are common parasites in domesticated cattle worldwide, including in Japan. We carried out a serological survey to detect C. parvum and N. caninum infection among cattle in the southern Kyushu region of Japan—including the small islands—by indirect enzyme-linked immunosorbent assay based on recombinant antigens. We found that total seropositivity in 570 Japanese black cattle was 96.3% for C. parvum and 18.4% for N. caninum. Although seroprevalence was correlated with cattle age, differences in the seroprevalence of C. parvum among age groups were not statistically significant. On the other hand, N. caninum seroprevalence increased with age, suggesting horizontal transmission through ingestion of food or water contaminated with oocysts. These findings underscore the importance of monitoring C. parvum and N. caninum in cattle and implementing measures to prevent the spread of infection to other livestock and to humans.     

Amblyomma americanum as a Bridging Vector for Human Infection with Francisella tularensis

The γ-proteobacterium Francisella tularensis causes seasonal tick-transmitted tularemia outbreaks in natural rabbit hosts and incidental infections in humans in the south-central United States. Although Dermacentor variabilis is considered a primary vector for F. tularensis, Amblyomma americanum is the most abundant tick species in this endemic region. A systematic study of F. tularensis colonization of A. americanum was undertaken to better understand its potential to serve as an overwintering reservoir for F. tularensis and as a bridging vector for human infections. Colony-reared A. americanum were artificially fed F. tularensis subspecies holarctica strain LVS via glass capillaries and colonization levels determined. Capillary-fed larva and nymph were initially infected with 10⁴ CFU/tick which declined prior to molting for both stages, but rebounded post-molting in nymphs and persisted in 53% at 10³ to 10⁸ CFU/nymph at 168 days post-capillary feeding (longest sampling time in the study). In contrast, only 18% of adults molted from colonized nymphs maintained LVS colonization at 10¹ to 10⁵ CFU/adult at 168 days post-capillary feeding (longest sampling time). For adults, LVS initially colonized the gut and disseminated to salivary glands by 24 h and had an ID₅₀ of <5CFU in mice. Francisella tularensis infected the ovaries of gravid females, but transmission to eggs was infrequent and transovarial transmission to hatched larvae was not observed. The prolonged persistence of F. tularensis in A. americanum nymphs supports A. americanum as an overwintering reservoir for F. tularensis from which seasonal epizootics may originate; however, although the rapid dissemination of F. tularensis from gut to salivary glands in adults A. americanum is compatible with intermittent feeding adult males acting as bridging vectors for incidental F. tularensis infections of humans, acquisition of F. tularensis by adults may be unlikely based on adult feeding preference for larger mammals which are not involved in maintenance of sylvatic tularemia.     

Factores asociados con la presencia de endoparásitos y ectoparásitos en perros domiciliados de la zona metropolitana de Toluca,México

Introduction:

Endoparasites and ectoparasites in dogs are of global distribution. The close relationship between dogs and man implies a risk for the transmission of zoonotic parasites. Therefore, it is necessary to determine the parasites hosted by dogs in specific areas and the factors associated with their presence.

Objectives:

To identify and to estimate the prevalence of endoparasites and ectoparasites in domiciled dogs in the Metropolitan area of Toluca, México, and the prevalence of D. caninum in fleas of the genus Ctenocephalides spp.

Materials and methods:

We collected samples from 402 domiciled dogs in four reference hospitals in the area in Toluca. We diagnosed endoparasites using direct smear, flotation, and sedimentation techniques and we performed the taxonomic identification of ectoparasites. Finally, the molecular diagnosis of D. caninum in fleas was made using the polymerase chain reaction technique (PCR).

Results:

A total of 37.2% of dogs were positive for endoparasites; the genera or species identified were Toxocara spp., Giardia spp., Ancylostoma spp., Cystoisospora spp., D. caninum, Taenia spp., and Trichuris vulpis; the prevalence of ectoparasites was 13.13%. We identified fleas of the species Ctenocephalides felis, Ctenocephalides canis; only one animal was parasitized with Rhipicephalus sanguineus and another one with Trichodectes canis; the prevalence of D. caninum in fleas was 9.5%.

Conclusion:

The prevalence of endoparasites was 37.2% while that of ectoparasites was 13.1%; this is the first analysis of endoparasites and ectoparasites conducted in the same population of dogs in México together with the molecular diagnosis of D. caninum in fleas.     

Development of competitive ELISA for neosporosis by employing immunoproteomics

In this study, proteomics was used to explore the antigenic proteins that are involved in cross-reactivity during serodiagnosis between Neospora caninum (N. caninum) and Toxoplasma gondii (T. gondii). Competitive enzyme-linked immunosorbent assay (C-ELISA) developed by proteomics shed a new light on the infection of N. caninum. Cross-reactivity of antigenic proteins between N. caninum and T. gondii tachyzoites was explored by using the conventional sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) (1-DE) and two-dimensional gel electrophoresis (2-DE) immunoblot. The proteins were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry. The protein expression patterns in the immunoblot profiles of N. caninum were similar to bovine, chicken, and rabbit anti-N. caninum serum, but they were not similar to rabbit anti-T. gondii serum. Band at 79 kDa, HSP70, and actin on immunoblot profiles reacted, in general, with bovine, chicken, and rabbit anti-N. caninum serum and also with rabbit anti-T. gondii serum, respectively. Whereas the band at 144 kDa, and NCDG-1 were detected on bovine, chicken, and rabbit anti-N. caninum immunoblot profiles, they were not observed on rabbit anti-T. gondii immunoblot profile. These specific antigenic proteins were recorded as species-specific proteins of N. caninum against T. gondii. Based on the proteome analysis, C-ELISA was developed to screen the cattle infected with N. caninum by using N. caninum tachyzoite lysate as a coating antigen and chicken anti-N. caninum immunoglobulin (Ig)Y as a competitor. C-ELISA was able to detect the antibody of N. caninum without cross-reactivity with T. gondii. Furthermore, it achieved a fine diagnostic performance in the cases of 162 bovine sera.     

Vertebrate-Aedes aegypti and Culex quinquefasciatus (Diptera)-arbovirus transmission networks: Non-human feeding revealed by meta-barcoding and next-generation sequencing

BackgroundAedes aegypti mosquito-borne viruses including Zika (ZIKV), dengue (DENV), yellow fever (YFV), and chikungunya (CHIKV) have emerged and re-emerged globally, resulting in an elevated burden of human disease. Aedes aegypti is found worldwide in tropical, sub-tropical, and temperate areas. The characterization of mosquito blood meals is essential to understand the transmission dynamics of mosquito-vectored pathogens.Methodology/principal findingsHere, we report Ae. aegypti and Culex quinquefasciatus host feeding patterns and arbovirus transmission in Northern Mexico using a metabarcoding-like approach with next-generation deep sequencing technology. A total of 145 Ae. aegypti yielded a blood meal analysis result with 107 (73.8%) for a single vertebrate species and 38 (26.2%) for two or more. Among the single host blood meals for Ae. aegypti, 28.0% were from humans, 54.2% from dogs, 16.8% from cats, and 1.0% from tortoises. Among those with more than one species present, 65.9% were from humans and dogs. For Cx. quinquefasciatus, 388 individuals yielded information with 326 (84%) being from a single host and 63 (16.2%) being from two or more hosts. Of the single species blood meals, 77.9% were from dogs, 6.1% from chickens, 3.1% from house sparrows, 2.4% from humans, while the remaining 10.5% derived from other 12 host species. Among those which had fed on more than one species, 11% were from dogs and humans, and 89% of other host species combinations. Forage ratio analysis revealed dog as the most over-utilized host by Ae. aegypti (= 4.3) and Cx. quinquefasciatus (= 5.6) and the human blood index at 39% and 4%, respectively. A total of 2,941 host-seeking female Ae. aegypti and 3,536 Cx. quinquefasciatus mosquitoes were collected in the surveyed area. Of these, 118 Ae. aegypti pools and 37 Cx. quinquefasciatus pools were screened for seven arboviruses (ZIKV, DENV 1–4, CHIKV, and West Nile virus (WNV)) using qRT-PCR and none were positive (point prevalence = 0%). The 95%-exact upper limit confidence interval was 0.07% and 0.17% for Ae. aegypti and Cx. quinquefasciatus, respectivelyConclusions/significanceThe low human blood feeding rate in Ae. aegypti, high rate of feeding on mammals by Cx. quinquefasciatus, and the potential risk to transmission dynamics of arboviruses in highly urbanized areas of Northern Mexico is discussed.