Person: Bezos Garrido, Javier
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First Name
Javier
Last Name
Bezos Garrido
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Veterinaria
Department
Sanidad Animal
Area
Sanidad Animal
Identifiers
31 results
Search Results
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Item Evaluation of a new enzyme-linked immunosorbent assay for the diagnosis of tuberculosis in goat milk(Research in Veterinary Science, 2020) Roy, A.; Infantes-Lorenzo, J.A.; Dominguez, M.; Moreno, I.; Pérez Sancho, Marta; García Benzaquén, Nerea; García-Seco Romero, María Teresa; Álvarez Sánchez, Julio; Romero Martínez, Beatriz; Gortázar, C.; Juan Ferré, Lucía De; Domínguez Rodríguez, Lucas José; Bezos Garrido, JavierCaprine tuberculosis (TB) is a zoonosis with sanitary and economic repercussions. Caprine TB control programs are based on a test and cull strategy using the intradermal tuberculin tests and slaughterhouse surveillance. However, this approach is not always feasible and may have a limited sensitivity under specific circumstances. In this study, performance of a new experimental test based on the P22 protein complex (P22 ELISA) was evaluated in two TB-infected herds using milk and serum samples and compared with cell-based diagnostic tests. Samples from a low (n = 62, herd 1) and a high (n = 52, herd 2) TB prevalence herd were selected. Moreover, bulk tank milk samples from both herds were analysed using the P22 ELISA. At the end of the study, a group of animals (n = 21) was euthanized and subjected to post-mortem analysis and bacteriological culture. Significant differences (p < .001) on the qualitative and quantitative (ODs) results were observed between herds using both serum and milk samples in the P22 ELISA. The correlation observed in the quantitative results obtained in serum and milk samples was very strong in animals from flock 2 (rs = 0.91) and moderate in animals from flock 1 (rs = 0.46). Among the slaughtered animals, the P22 ELISA detected a higher proportion of lesion-culture positive animals than cell-based diagnostic tests (61.9 and 66.7% using milk and serum samples, respectively). The P22 ELISA using milk samples demonstrated a similar sensitivity compared with serum samples, suggesting it might be a valuable test for TB control in dairy goats.Item Comparison of four different culture media for isolation and growth of type II and type I/III Mycobacterium avium subsp. paratuberculosis strains isolated from cattle and goats(Applied and Environmental Microbiology, 2006) Juan Ferré, Lucía De; Álvarez Sánchez, Julio; Romero Martínez, Beatriz; Bezos Garrido, Javier; Castellanos, Elena; Aranaz Martín, Alicia; Mateos García, Ana Isabel; Domínguez Rodríguez, Lucas JoséCulture is considered the definitive technique for Johne's disease diagnosis, and it is essential for later applications of certain molecular typing techniques. In this study, we have tested four solid media (Herrold's egg yolk medium [HEYM] with sodium pyruvate and mycobactin [HEYMm-SP], HEYM with mycobactin and without sodium pyruvate [HEYMm], Middlebrook 7H11 with mycobactin [Mm], and Löwenstein-Jensen with mycobactin [LJm]) for isolation of Mycobacterium avium subsp. paratuberculosis strains in 319 tissue samples from cattle herds and goat flocks. We have shown that each of the two main groups of M. avium subsp. paratuberculosis (type II and type I/III) has different requirements for growth in the culture media studied. The recommended solid media for isolation of type I/III strains are LJm and Mm, since the combination of both media allowed the recovery of all these strains. The most widespread culture medium, HEYM, is not suitable for the isolation of this group of M. avium subsp. paratuberculosis strains. Regarding the type II strains, HEYMm-SP was the medium where more strains were isolated, but the other three media are also needed in order to recover all type II strains. The incubation period is also related to the strain type. In conclusion, because the type of strain cannot be known in advance of culture, coupled with the fact that cattle and goats can be infected with both groups of strains, we recommend the use of the four solid media and the prolongation of the incubation period to more than 6 months to detect paratuberculous herds/flocks and to determine the true prevalence of the infection.Item Factors affecting the performance of P22 ELISA for the diagnosis of caprine tuberculosis in milk samples(Research in Veterinary Science, 2022) Ortega Becerril, José Antonio; Infantes Lorenzo, José Antonio; Roy, A.; Juan Ferré, Lucía De; Romero Martínez, Beatriz; Moreno, I.; Domínguez, M.; Bezos Garrido, Javier; Domínguez Rodríguez, Lucas JoséCaprine tuberculosis (TB) is a zoonosis caused by members of the Mycobacterium tuberculosis complex (MTBC). Caprine TB eradication programmes are based mainly on intradermal tuberculin tests and slaughterhouse surveillance. However, the use of serological test has been extended as a potential diagnostic tool in goats through the use of serum, plasma, or even milk samples. Milk production and the antibodies (Ab) present in milk can vary depending on several circumstances. In the present study, different factors that may affect the performance of humoral TB diagnosis were analysed using goat milk samples: 1) lactation stage, 2) a recent previous skin test (booster effect) and 3) the effect of freeze-thaw cycles on milk samples preserved with azidiol. TB-infected animals (n = 44) were selected to evaluate the evolution of the Ab levels during the 6-month lactation period, along with its potential effect on the P22 ELISA results. In general, no significant changes (p = 0.079) were observed throughout the study as regards Ab levels in milk samples between consecutive analysis although the reactivity to P22 ELISA decreased when samplings were performed at the last two months of the lactation. Regarding the booster effect, the quantitative results showed a significant variation (p < 0.001) for both milk and serum samples when serological tests were carried out 15 days after the skin test. Finally, there were no significant differences (p = 0.99) in the P22 ELISA results when using milk samples preserved with azidiol that had undergone freeze-thaw cycles.Item Assessment of the sensitivity of the bovine tuberculosis eradication program in a high prevalence region of Spain using scenario tree modeling(Preventive Veterinary Medicine, 2019) De la Cruz, María Luisa; Pozo Piñol, Pilar; Grau, Anna; Nacar, Jesús; Bezos Garrido, Javier; Pérez, Andrés; Domínguez Rodríguez, Lucas José; Sáez, José Luís; Mínguez-González, Olga; Juan Ferré, Lucía De; Álvarez Sánchez, JulioIn spite of the efforts invested to eradicate bovine tuberculosis (bTB) in cattle, the disease is still present in several developed countries, including Spain. Among the factors related with disease persistence in high prevalence areas, the lack of sensitivity of the screening test for detection of infected herds [single intradermal tuberculin (SIT) test] can play a major role. Here, a scenario tree model mimicking the diagnostic test scheme in place in the region of Castilla y Leon (Spain) was developed to estimate the probability of detecting bTB in an infected-non detected officially tuberculosis free (OTF) herd (herd sensitivity, HSe). In order to do so the probability of detecting at least one positive animal in the SIT test with/without post-mortem (detection of lesions and culture) confirmation in an infected herd was estimated using Monte Carlo simulation through @ RISK (Palisade Co, NY, USA). Uncertainty on the accuracy of the diagnostic tests was introduced in the model using distributions based on the literature. The performance of the model was evaluated by comparing the predicted number of SIT/post-mortem positive animals in infected herds with those observed in newly detected bTB-infected herds in the region in 2011–2015. The estimated HSe of the SIT test was 76.2% (95% probability interval: 19.8–97.6). According to the model, bTB infection would be then confirmed through culture in 65.3% (95% PI: 50.0–82.3) of the herds detected through the SIT test, so that overall the proportion of infected-non detected OTF herds in which the infection could be confirmed after the initial SIT test was 49.6% (95% PI: 9.75–80.3). The predicted HSe of both SIT test and culture was directly correlated with herd size. Results from the model suggest a moderate but highly variable HSe of the current surveillance system in place for bTB detection in OTF herds located in high prevalence areas, that could be maximized by performing multiple tests within a year as indicated in the Spanish eradication program (with a median SIT HSe of 87% when two consecutive tests were considered). In addition, these results highlight the usefulness of performing subsequent SIT tests to rule out infection in SIT-positive herds even when the causative agent cannot be isolated.Item Effect of paratuberculosis on the diagnosis of bovine tuberculosis in a cattle herd with a mixed infection using interferon-gamma detection assay(Veterinary Microbiology, 2009) Álvarez Sánchez, Julio; Juan Ferré, Lucía De; Bezos Garrido, Javier; Romero Martínez, Beatriz; Sáez, Jose Luis; Marqués, Sergio; Domínguez, Concepción; Mínguez, Olga; Fernández-Mardomingo, Baudilio; Mateos García, Ana Isabel; Domínguez Rodríguez, Lucas José; Aranaz Martín, AliciaInterferon-gamma (IFN-gamma) detection assay is being applied as an ancillary test to tuberculin tests in the diagnosis of bovine tuberculosis to detect the maximum number of infected animals. Among possible factors influencing the performance of tuberculosis-diagnostic tests, paratuberculosis, a widespread disease in Spain and other European countries, has been pointed out as a cause of false positive reactions. Still, its effect on the sensitivity of these tests in cattle has yet to be fully characterized. The impact of paratuberculosis in the apparent sensitivity of IFN-gamma assay was studied in a bullfighting cattle herd with a mixed tuberculosis-paratuberculosis infection, using culture of Mycobacterium bovis and Mycobacterium avium paratuberculosis as the gold standard to determine the infection status of every animal. A total of 218 animals were slaughtered and sampled for bacteriology after blood sampling. IFN-gamma assay showed a lower apparent sensitivity in animals with a mixed infection (50%) compared to all animals suffering tuberculosis (78.3%). This finding indicates that the presence of paratuberculosis in tuberculosis-infected herds could imply a serious impairment in the sensitivity of IFN-gamma detection test.Item Genetic diversity of Mycobacterium avium isolates recovered from clinical samples and from the environment: molecular characterization for diagnostic purposes(Journal of clinical microbiology, 2008) Álvarez Sánchez, Julio; Gómez García, Ignacio; Aranaz Martín, Alicia; Bezos Garrido, Javier; Romero Martínez, Beatriz; Juan Ferré, Lucía De; Mateos García, Ana Isabel; Gómez Mampaso, Enrique; Domínguez Rodríguez, Lucas JoséIsolation of Mycobacterium avium complex (MAC) organisms from clinical samples may occur in patients without clinical disease, making the interpretation of results difficult. The clinical relevance of MAC isolates from different types of clinical samples (n = 47) from 39 patients in different sections of a hospital was assessed by comparison with environmental isolates (n = 17) from the hospital. Various methods for identification and typing (commercial probes, phenotypic characteristics, PCR for detection of IS1245 and IS901, sequencing of the hsp65 gene, and pulsed-field gel electrophoresis) were evaluated. The same strain was found in all the environmental isolates, 21 out of 23 (91.3%) of the isolates cultured from urine samples, and 5 out of 19 (26.3%) isolates from respiratory specimens. This strain did not cause disease in the patients. Testing best characterized the strain as M. avium subsp. hominissuis, with the unusual feature that 81.4% of these isolates lacked the IS1245 element. Contamination of certain clinical samples with an environmental strain was the most likely event; therefore, characterization of the environmental mycobacteria present in health care facilities should be performed to discard false-positive isolations in nonsterile samples, mainly urine samples. Molecular techniques applied in this study demonstrated their usefulness for this purpose.Item Evaluation of the Effect of a Recent Comparative Intradermal Tuberculin Test on the Humoral Diagnosis of Paratuberculosis Using Serum and Milk Samples from Goats(Veterinary Science, 2024) Velasco, Carlos; Ortega, Javier; Gómez-Buendía, Alberto; Grau, Anna; López, Marisol; Álvarez Sánchez, Julio; Romero Martínez, Beatriz; Juan Ferré, Lucía De; Bezos Garrido, JavierParatuberculosis (PTB) and tuberculosis (TB) are two mycobacterial diseases with a severe economic and health impact on domestic ruminants. The ante mortem diagnosis of PTB is hampered, among other factors, by the limited sensitivity of all the available diagnostic techniques. Since TB-infected goats subjected to the comparative intradermal tuberculin test (CITT) may experience a booster effect on their antibody titer and a potential enhancement to the sensitivity of humoral techniques for tuberculosis, in the present study we aimed to evaluate this diagnostic strategy on the humoral diagnosis of PTB in serum and milk samples collected from a caprine herd that was TB free and PTB infected. The results from 120 goats indicated a significant increase (p < 0.001) in the quantitative response detected using an ELISA technique, conducted using serum and milk samples taken 15 and 30 days after performing a CITT (day 0 of the study); although, it did not translate into a significant increase in the number of reactors during any of the testing events (0, 3,15, 30 and 60 days post-CITT). Additionally, the number of ELISA-positive animals was higher for the serum versus the milk samples at both 15 and 30 days post-CITT. The increase in the quantitative ELISA result suggested a diagnostic strategy that maximizes ELISA sensitivity, mainly using serum samples, in PTB-infected herds; although, it may depend on individual differences and the interpretation criteria.Item Assessment of diagnostic tools for eradication of bovine tuberculosis in cattle co-infected with Mycobacterium bovis and M. avium subsp. paratuberculosis(Veterinary Research, 2006) Aranaz Martín, Alicia; Juan Ferré, Lucía De; Bezos Garrido, Javier; Álvarez Sánchez, Julio; Romero Martínez, Beatriz; Lozano, Francisco; Paramio, José L.; López-Sánchez, Jesús; Mateos García, Ana Isabel; Domínguez Rodríguez, Lucas JoséThe intradermal tuberculin (IDTB) test and the interferon-gamma (IFN-$\gamma$) assay are used worldwide for detection of bovine tuberculosis in cattle, but little is known about the effect of co-infecting agents on the performance of these diagnostic tests. This report describes a field trial conducted in a cattle herd with dual infection (bovine tuberculosis and paratuberculosis) during 3.5 years. It has been based on a strategic approach encompassing serial parallel testing (comparative IDTB test, the IFN-$\gamma$ assay and serology of paratuberculosis) that was repeated 8 times over the period, and segregation of animals into two herds. The IDTB test detected 65.2% and the IFN-$\gamma$ test detected 69.6% of the Mycobacterium bovis culture-positive cattle. However, the IDTB test performed better during the first part of the trial, while the IFN-$\gamma$ test was the only method that detected infected animals during the following three samplings. The number of false positive reactors with the IDTB and/or the IFN-$\gamma$ tests was remarkably high compared to other reports, and could be caused by cross-reactivity with M. avium subsp. paratuberculosis. Also, the M. bovis isolates from cattle and wildlife from the same property were characterised using molecular techniques to disclose an epidemiological link. The IDTB test may not be appropriate to eradicate bovine tuberculosis in herds with dual mycobacterial infections. This report highlights the need to use several diagnostic techniques for the accurate detection of M. bovis infected animals in these herds.Item Use of Whole-Genome Sequencing to Unravel the Genetic Diversity of a Prevalent Mycobacterium bovis Spoligotype in a Multi-Host Scenario in Spain(Frontiers in Microbiology, 2022) Pozo Piñol, Pilar; Lorente Leal, Víctor; Robbe-Austerman, Suelee; Hicks, Jessica; Stuber, Tod; Bezos Garrido, Javier; Juan Ferré, Lucía De; Sáez-Lorente, José Luis; Romero Martínez, Beatriz; Álvarez Sánchez, JulioDespite the efforts invested in the eradication of bovine tuberculosis in Spain, herd prevalence has remained constant in the country during the last 15 years (~1.5–1.9%) due to a combination of epidemiological factors impairing disease control, including between-species transmission. Here, our aim was to investigate the molecular diversity of Mycobacterium bovis isolates belonging to the highly prevalent SB0339 spoligotype in the cattle-wildlife interface in different regions of Spain using whole-genome sequencing (WGS). Genomic data of 136 M. bovis isolates recovered from different animal species (cattle, wild boar, fallow deer, and red deer) and locations between 2005 and 2018 were analyzed to investigate between- and within-species transmission, as well as within-herds. All sequenced isolates differed by 49–88 single nucleotide polymorphisms from their most recent common ancestor. Genetic heterogeneity was geographic rather than host species-specific, as isolates recovered from both cattle and wildlife from a given region were more closely related compared to isolates from the same species but geographically distant. In fact, a strong association between the geographic and the genetic distances separating pairs of M. bovis isolates was found, with a significantly stronger effect when cattle isolates were compared with wildlife or cattle-wildlife isolates in Spain. The same results were obtained in Madrid, the region with the largest number of sequenced isolates, but no differences depending on the host were observed. Within-herd genetic diversity was limited despite the considerable time elapsed between isolations. The detection of closely related strains in different hosts demonstrates the complex between-host transmission dynamics present in endemic areas in Spain. In conclusion, WGS results a valuable tool to track bTB infection at a high resolution and may contribute to achieve its eradication in Spain.Item Effect of a recent intradermal test on the specificity of P22 ELISA for the diagnosis of caprine tuberculosis(Frontiers in Veterinary Science, 2024) Romero Martínez, Beatriz; Juan Ferré, Lucía De; Domínguez Rodríguez, Lucas José; Álvarez Sánchez, Julio; Bezos Garrido, Javier; Velasco, Carlos; Ortega, Javier; Domínguez, Mercedes; Ricón, Jaime; Moreno, InmaculadaCaprine tuberculosis (TB) is a zoonotic disease caused by members of the Mycobacterium tuberculosis complex. TB eradication programs in goats are based on the single and comparative intradermal tuberculin tests (SITT and CITT, respectively). Antibody-based diagnostic techniques have emerged as potential diagnostic tools for TB. P22 ELISA has been previously evaluated using samples collected after the intradermal tuberculin tests to maximize the sensitivity, a phenomenon known as booster effect. However, there is no information available on whether the use of this diagnostic strategy could lead to a decrease of its specificity (Sp). The aim of the present study was to elucidate the interference effect of a recent CITT on the Sp of the P22 ELISA in serum and milk samples collected at different times after the CITT from a TB-free herd (n = 113). The number of reactors to P22 ELISA was significantly higher (p < 0.01) on serum samples collected 15 days post-CITT compared to day 0, showing a decrease in Sp from 99.1% (95% CI; 95.2–99.8%) to 88.5% (95% CI; 81.3–93.2%). The number of reactors and the quantitative values of P22 ELISA were significantly higher (p < 0.01) in serum samples compared to milk. No significant (p > 0.05) changes in the Sp of the P22 ELISA were observed throughout the different time samplings using milk No significant (p > 0.05) changes were observed on days 30 and 60 post-CITT. In conclusion, the booster effect strategy may significantly decrease the Sp of P22 ELISA in TB-free herds when serum samples are used but not when milk is tested.