Vetgenomics

La Línea de la Concepción, Spain

Vetgenomics

La Línea de la Concepción, Spain

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Rivas L.,Catalan Institute of Nanoscience and Nanotechnology | Rivas L.,Autonomous University of Barcelona | de la Escosura-Muniz A.,Catalan Institute of Nanoscience and Nanotechnology | Serrano L.,Vetgenomics | And 6 more authors.
Nano Research | Year: 2015

A novel triple lines lateral-flow assay (LFA) with enhanced sensitivity for the detection of Leishmania infantum DNA in dog blood samples was designed and successfully applied. The enhanced LFA methodology takes advantage of the gold nanoparticle tags (AuNPs) conjugated to polyclonal secondary antibodies, which recognize anti-FITC antibodies. The polyclonal nature of the secondary antibodies allows for multiple binding to primary antibodies, leading to enhanced AuNP plasmonics signal. Furthermore, endogenous control consisting of the amplified dog 18S rRNA gene was introduced to avoid false negatives. Using this strategy, 0.038 spiked Leishmania parasites per DNA amplification reaction (1 parasite/100 μL of DNA sample) were detected. Detection limit of LFA was found to be lower than that of the conventional techniques. In summary, our novel LFA design is a universal and simple sensing alternative that can be extended to several other biosensing scenarios. [Figure not available: see fulltext.] © 2015, Tsinghua University Press and Springer-Verlag Berlin Heidelberg.


Pennisi M.-G.,Messina University | Persichetti M.-F.,Messina University | Serrano L.,Vetgenomics | Altet L.,Vetgenomics | And 3 more authors.
Parasites and Vectors | Year: 2015

Background: Limited information is available about the species of ticks infesting the cat and the pathogens that they harbor. The aims of the present study were to identify the species of ticks removed from cats living in Sicily and Calabria (Italy) and to detect DNA of vector-borne pathogens in the same ticks. Findings: Morphological identification of 132 adult ticks collected throughout the year from cats was carried out. Real-time PCRs for Hepatozoon felis, Piroplasmid, Ehrlichia/Anaplasma spp., Rickettsia spp., Bartonella spp., Mycoplasma spp. and Leishmania infantum were performed from each individual tick. Ticks belonging to Rhipicephalus (R. sanguineus sensu lato, R. pusillus) and Ixodes (I. ricinus, I. ventalloi) genera were identified. Ixodes ventalloi was the most frequently found tick species (47 %). The positivity rate to at least one pathogen was 14.4 % (19/132 ticks). Leishmania infantum, Rickettsia spp. (R. monacensis and R. helvetica), Bartonella spp. (B. clarridgeiae), Piroplasmid (Babesia vogeli), and Ehrlichia/Anaplasma spp. (E. canis) DNAs were amplified in 8.3, 5.3, 1.5, 0.75 and 0.75 % of ticks, respectively. Hepatozoon felis, Anaplasma spp. and hemotropic Mycoplasma spp. DNAs were not detected. Four (21.1 %) out of nineteen positive ticks were co-infected. Conclusions: This study provides novel data about ticks infesting cats and the DNA of pathogens that they harbor. In Southern Italy, anti-tick prophylaxis should be implemented throughout the year in cats without neglecting winter time. © 2015 Pennisi et al.


PubMed | Vetgenomics, Catalan Institute of Nanoscience and Nanotechnology and Autonomous University of Barcelona
Type: Journal Article | Journal: Small (Weinheim an der Bergstrasse, Germany) | Year: 2016

A novel methodology for the isothermal amplification of Leishmania DNA using labeled primers combined with the advantages of magnetic purification/preconcentration and the use of gold nanoparticle (AuNP) tags for the sensitive electrochemical detection of such amplified DNA is developed. Primers labeled with AuNPs and magnetic beads (MBs) are used for the first time for the isothermal amplification reaction, being the amplified product ready for the electrochemical detection. The electrocatalytic activity of the AuNP tags toward the hydrogen evolution reaction allows the rapid quantification of the DNA on screen-printed carbon electrodes. Amplified products from the blood of dogs with Leishmania (positive samples) are discriminated from those of healthy dogs (blank samples). Quantitative studies demonstrate that the optimized method allows us to detect less than one parasite per microliter of blood (8 10(-3) parasites in the isothermal amplification reaction). This pioneering approach is much more sensitive than traditional methods based on real-time polymerase chain reaction (PCR), and is also more rapid, cheap, and user-friendly.


PubMed | Instituto Zooprofilattico Sperimentale della Sicilia, Veterinary practitioner, Vetgenomics, Messina University and Autonomous University of Barcelona
Type: | Journal: Parasites & vectors | Year: 2015

Limited information is available about the species of ticks infesting the cat and the pathogens that they harbor. The aims of the present study were to identify the species of ticks removed from cats living in Sicily and Calabria (Italy) and to detect DNA of vector-borne pathogens in the same ticks.Morphological identification of 132 adult ticks collected throughout the year from cats was carried out. Real-time PCRs for Hepatozoon felis, Piroplasmid, Ehrlichia/Anaplasma spp., Rickettsia spp., Bartonella spp., Mycoplasma spp. and Leishmania infantum were performed from each individual tick. Ticks belonging to Rhipicephalus (R. sanguineus sensu lato, R. pusillus) and Ixodes (I. ricinus, I. ventalloi) genera were identified. Ixodes ventalloi was the most frequently found tick species (47%). The positivity rate to at least one pathogen was 14.4% (19/132 ticks). Leishmania infantum, Rickettsia spp. (R. monacensis and R. helvetica), Bartonella spp. (B. clarridgeiae), Piroplasmid (Babesia vogeli), and Ehrlichia/Anaplasma spp. (E. canis) DNAs were amplified in 8.3, 5.3, 1.5, 0.75 and 0.75% of ticks, respectively. Hepatozoon felis, Anaplasma spp. and hemotropic Mycoplasma spp. DNAs were not detected. Four (21.1%) out of nineteen positive ticks were co-infected.This study provides novel data about ticks infesting cats and the DNA of pathogens that they harbor. In Southern Italy, anti-tick prophylaxis should be implemented throughout the year in cats without neglecting winter time.


News Article | November 27, 2015
Site: www.nanotech-now.com

Abstract: In an article published in Small, researchers successfully applied a new qualitative and quantitative method for the detection of a DNA sequence characteristic of Leishmania infantum kinetoplast, a frequent parasite in veterinary that affects humans too. The work was led from the Catalan Institute of Nanoscience and Nanotechnology (ICN2), a research Center placed in the Campus of the Autonomous University of Barcelona (UAB) in Bellaterra, and the UAB Spin Off company Vetgenomics. This work was coordinated at ICN2 by ICREA Prof Arben Merkoçi, Leader of the ICN2 Nanobioelectronics & Biosensors Group, and Dr. Alfredo de la Escosura-Muñiz (first author of the article) with collaboration of Luis Pires, PhD student from the same group. The work was carried out in collaboration with Prof. Armand Sanchez, Dr. Olga Francino, Dr. Laura Altet and Lorena Serrano from Vetgenomics. Their results are part of the bio-applications defined in the ICN2 Severo Ochoa Program "Devices for Social Challenges". The present work has been published within the POC4PETS European Project, coordinated by the Alma Mater Studiorum - Università di Bologna (Italy) and aimed to improving the speed and accuracy of current diagnostics for veterinary pathogens. Overcoming the classical PCR technique The Polymerase Chain Reaction (PCR) is today's standard method to identify the presence of a specific DNA sequence in a sample. The PCR uses enzymes and two primers, strands of short nucleic acid sequences that serve as a starting point for DNA copy. When the detection is positive, this technique produces millions of copies of the problem sequence to facilitate its detection. This DNA amplification involves precise thermal changes (thermocycling) and sophisticate and expensive equipment which are overcome by an alternative approach called isothermal amplification, performed at constant temperature. In this context, the authors of the article present a novel design of isothermal amplification using primers labelled with both gold nanoparticles and magnetic microbeads. The amplified product carries both labels allowing a rapid purification and quantification. The magnetic properties of the first primer facilitate the purification/preconcentration of the amplified DNA through magnetic separation methods. On the other hand, the gold nanoparticles are easily quantified by simple electrocatalytic detection methods. Thus, the use of primers labelled with gold nanoparticles and magnetic microbeads turns isothermal amplification into a faster and easier qualitative and quantitative diagnostic method. Nanoparticles for Leishmania detection and other point-of-care tests This approach was successfully applied for the detection of a DNA sequence characteristic of Leishmania infantum kinetoplast, a parasite responsible of a disease in domestic dogs, wild canids and humans. The electrochemical method exhibited a good reproducibility and sensitivity. Furthermore, amplified DNA from dogs without Leishmania was perfectly discriminated, demonstrating the specificity of both the amplification procedure and the electrochemical detection. In fact, the performance of the proposed approach is better than the obtained with other point-of-care tests for Leishmania detection, offering also a quantitative tool for parasite determination. This method represents a universal methodology that could be applied for any isothermal DNA amplification design. The technology presented in this article is under patenting process (EP14382266.6/P10398EP00 (09/07/14); PCT/EP2015/065742 (09/07/15)). ### About POC4PETS: Veterinary diagnostics is a key tool in the prevention and control of infectious diseases in animals. It is increasingly recognized that there is the urgent need for innovation in the animal infectious diseases testing field, bringing to new market applications in the development of specific, rapid and efficiently validated diagnostic tests. The European FP7 project Point of Care Diagnostics for rapid and cheap pathogen detection of companion animals (POC4PETS) aims to apply the most promising new technologies to improving both the speed and accuracy of the current diagnostics for veterinary pathogens of companion animal species. ICN2's Nanobioelectronics & Biosensors Group, led by ICREA Research Professor Arben Merkoçi, is one of the POC4PETS partners. Their research focuses in the development of novel nanotechnology and nanomaterials based sensors for DNA, protein and cells detection. The project is particularly focused in filling the gaps of diagnostics availability and enhancing technology transfer to increase competitiveness of the Veterinary Diagnostic industry. Vetgenomics is a SME spin-off of the UAB (Universitat Autònoma de Barcelona) founded in 2010 and devoted to molecular diagnostic in companion animals. The research activity is mainly focused in the animal genomics and veterinary genetic diagnostic fields. The main goal is becoming a company focused in innovation and with the flexibility to adapt technology to customer needs, increasing the added value of their products and becoming a partner of choice in R+D+i. For more information, please click If you have a comment, please us. Issuers of news releases, not 7th Wave, Inc. or Nanotechnology Now, are solely responsible for the accuracy of the content.


PubMed | Vetgenomics, Messina University, Autonomous University of Barcelona and Instituto Zooprofilattico Sperimentale della Sicilia Adelmo Mirri
Type: Journal Article | Journal: Parasites & vectors | Year: 2016

Vector-borne pathogens are the subject of several investigations due to the zoonotic concern of some of them. However, limited data are available about the simultaneous presence of these pathogens in cats and their ectoparasites. The aim of the present study was to define the species of ectoparasites found on cats as well as to investigate vector-borne pathogens in cats and their ectoparasites in southern Italy.Blood from 42 cats and fleas or flea pools (n=28) and ticks (n=73) collected from them were investigated by quantitative PCR for the detection of vector-borne pathogens. Feline serum samples were tested by IFAT to detect IgG antibodies against Leishmania infantum, Bartonella henselae, Rickettsia conorii, Rickettsia felis, Rickettsia typhi, Babesia microti, Ehrlichia canis and Anaplasma phagocytophilum antigens.Only one flea species (Ctenocephalides felis) and four tick species belonging to the genera Rhipicephalus and Ixodes were identified on cats from southern Italy. Molecular evidence of Bartonella spp., Rickettsia spp., hemoplasmas, Babesia vogeli and L. infantum was found in ectoparasites (fleas and/or ticks) while DNA from Hepatozoon felis and Ehrlichia/Anaplasma spp. was not detected. Likewise, DNAs from Bartonella, hemoplasma and Leishmania were the only pathogens amplified from feline blood samples. Cats had also antibodies against all the investigated pathogens with the exception of Rickettsia typhi. Agreement between serological and molecular results in individual cats and their ectoparasites was not found. The only exception was for Bartonella with a fair to moderate agreement between individual cats and their ectoparasites. Bartonella clarridgeiae was the species most frequently found in cats and their fleas followed by B. henselae.In conclusion, cats harboring ticks and fleas are frequently exposed to vector-borne pathogens. Furthermore, ticks and fleas harbored by cats frequently carry pathogens of zoonotic concern therefore appropriate feline ectoparasiticide preventative treatments should be used in cats.


PubMed | El Mirador del Rosario P5, Oklahoma State University, Health & Biotechnology SaBio, Major University and 2 more.
Type: Journal Article | Journal: Ticks and tick-borne diseases | Year: 2016

Urbanization of natural areas is considered one of the causes of the current apparent emergence of infectious diseases. Carnivores are among the species that adapt well to urban and periurban environments, facilitating cross-species disease transmission with domestic dogs and cats, and potentially with their owners. The prevalence of vector-borne pathogens (VBP) of zoonotic and veterinary interest was studied in sympatric wild and domestic carnivores into Barcelona Metropolitan Area (NE Spain). Blood or spleen samples from 130 animals, including 34 common genets (Genetta genetta), 12 red foxes (Vulpes vulpes), 10 stone martens (Martes foina), three Eurasian badgers (Meles meles), 34 free-roaming domestic cats and 37 dogs with outdoor access, were collected either in protected or adjacent residential areas. A total of 309 ticks (chiefly Rhipicephalus turanicus) were collected on these animals. The samples were analyzed with a battery of PCR assays targeting the DNA of Rickettsia spp., Anaplasmataceae, Coxiella burnetii, Bartonella spp., and Piroplasmida, and the amplicons were sequenced. The fox showed the highest prevalence (58%) and diversity of VBP (four pathogens), whereas none of the dogs were infected. Bartonella spp. (including B. clarridgeiae, B. henselae, and B. rochalimae) was the most prevalent pathogen. Infection of wild carnivores with Ehrlichia canis, C. burnetii, Theileria annae and Babesia vogeli was also confirmed, with some cases of coinfection observed. The presence of DNA of T. annae and B. vogeli was also confirmed in tick pools from four species of wild carnivores, supporting their role in piroplasmid life-cycle. By the sequencing of several target genes, DNA of Rickettsia massiliae was confirmed in 17 pools of Rh. turanicus, Rh. sanguineous, and Rh. pusillus from five different species, and Rickettsia conorii in one pool of Rh. sanguineous from a dog. None of the hosts from which these ticks were collected was infected by Rickettsia. Although carnivores may not be reservoir hosts for zoonotic Rickettsia, they can have an important role as mechanical dispersers of infected ticks.


Millan J.,Andrés Bello University | Chirife A.D.,El Mirador del Rosario P5 | Altet L.,Vetgenomics
Veterinary Quarterly | Year: 2015

Background: The role of wildlife in the epidemiology of leishmaniosis in under debate, and determining whether infection with Leishmania infantum causes illness in wild carnivores is important to determine its potential role as a reservoir. Objectives: To provide for the first time serum biochemistry reference values for the common genet (Genetta genetta), and to determine variations associated with L. infantum infection. Methods: Twenty-five serum biochemistry parameters were determined in 22 wild-caught genets. Blood samples were analyzed for L. infantum DNA by means of real-time polymerase chain reaction (PCR). Results: Two female genets were positive for L. infantum DNA but did not show any external clinical sign upon physical examination. Among other variations in the biochemistry values of these genets, one presented a higher concentration of gamma-globulins and cholesterol, whereas the other genet presented increased creatinine, bilirubin, and chloride levels when compared to uninfected females. Sex-related differences in some parameters were also reported. Conclusion: Infection with L. infantum may sometimes be accompanied by abnormal serum biochemistry in wild carnivores. Clinical importance: Clinical disease may occur in L. infantum-infected wild carnivores. This has implications in the epidemiology of leishmaniosis. In addition, the data provided here would also be useful as reference values for researchers or rehabilitators working with the common genet. © 2015, © 2014 Taylor & Francis.


Solano-Gallego L.,Autonomous University of Barcelona | Di Filippo L.,Autonomous University of Barcelona | Ordeix L.,Autonomous University of Barcelona | Planellas M.,Autonomous University of Barcelona | And 4 more authors.
Parasites and Vectors | Year: 2016

Background: Leishmania infantum-specific antibodies are used extensively for the diagnosis and monitoring of treatment in canine leishmaniosis. Different views have been described for the measurement of L. infantum antibody levels for the monitoring of anti-leishmanial treatment. In addition, molecular techniques using blood are frequently employed in the clinical setting. However, there are not enough studies to prove the usefulness of PCR in diagnosis, treatment monitoring and in assessing the prognosis of the disease. The objectives of this study were to evaluate L. infantum-specific antibodies and blood parasitemia at the time of diagnosis and during treatment and to correlate these with the dog's clinical status. Methods: Thirty-seven dogs were diagnosed and followed-up during treatment (days 30, 180 and 365). The treatment protocol consisted of a combination of meglumine antimoniate for one month and allopurinol for at least one year. Leishmania infantum-specific antibodies and blood parasitemia were assessed by an end point sera dilution ELISA and by real-time PCR, respectively. Results: The majority of dogs were classified as LeishVet stage II (moderate disease) at the time of diagnosis (86 %) and the rest as stage III. Results showed variable levels of specific antibodies at the time of diagnosis [median ± interquartile range (IQR): 1372 ± 8803 ELISA units (EU)]. Twenty-three seropositive dogs (64 %) were detected as PCR-positive at the time of diagnosis. Interestingly, a rapid significant antibody level reduction was observed by day 30 of treatment (median ± IQR: 604 ± 2168 EU). A continuing significant decrease of specific antibodies was also found at days 180 (median ± IQR: 201 ± 676 EU) and 365 (median ± IQR: 133 ± 329 EU) in association with clinical improvement. A significant blood parasitemia reduction was also observed at all time points studied. Mean parasites/ml ± SD were 19.4 ± 79.1 on day 0, 2.2 ± 11.7 on day 30, 0.9 ± 2.9 on day 180, and 0.3 ± 0.7 on day 365. Conclusions: This study reports a significant reduction of L. infantum antibodies measured by an end point sera dilution ELISA method after 30 days of treatment associated with clinical improvement. A low proportion of sick dogs with moderate disease were negative by blood real-time PCR at the time of diagnosis. © 2016 Solano-Gallego et al.


PubMed | Vetgenomics and Autonomous University of Barcelona
Type: Journal Article | Journal: Parasites & vectors | Year: 2016

Leishmania infantum-specific antibodies are used extensively for the diagnosis and monitoring of treatment in canine leishmaniosis. Different views have been described for the measurement of L. infantum antibody levels for the monitoring of anti-leishmanial treatment. In addition, molecular techniques using blood are frequently employed in the clinical setting. However, there are not enough studies to prove the usefulness of PCR in diagnosis, treatment monitoring and in assessing the prognosis of the disease. The objectives of this study were to evaluate L. infantum-specific antibodies and blood parasitemia at the time of diagnosis and during treatment and to correlate these with the dogs clinical status.Thirty-seven dogs were diagnosed and followed-up during treatment (days 30, 180 and 365). The treatment protocol consisted of a combination of meglumine antimoniate for one month and allopurinol for at least one year. Leishmania infantum-specific antibodies and blood parasitemia were assessed by an end point sera dilution ELISA and by real-time PCR, respectively.The majority of dogs were classified as LeishVet stage II (moderate disease) at the time of diagnosis (86%) and the rest as stage III. Results showed variable levels of specific antibodies at the time of diagnosis [medianinterquartile range (IQR): 13728803 ELISA units (EU)]. Twenty-three seropositive dogs (64%) were detected as PCR-positive at the time of diagnosis. Interestingly, a rapid significant antibody level reduction was observed by day 30 of treatment (medianIQR: 6042168 EU). A continuing significant decrease of specific antibodies was also found at days 180 (medianIQR: 201676 EU) and 365 (medianIQR: 133329 EU) in association with clinical improvement. A significant blood parasitemia reduction was also observed at all time points studied. Mean parasites/mlSD were 19.479.1 on day 0, 2.211.7 on day 30, 0.92.9 on day 180, and 0.30.7 on day 365.This study reports a significant reduction of L. infantum antibodies measured by an end point sera dilution ELISA method after 30days of treatment associated with clinical improvement. A low proportion of sick dogs with moderate disease were negative by blood real-time PCR at the time of diagnosis.

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