Conjugated linoleic acid modifies transcriptional cytokine profile and induces early specific secretory IgA response in Giardia lamblia infected mice.

Objectives: Adaptive immunity is crucial in controlling Giardia lamblia infection in the intestinal mucosa, and some dietary lipids may improve mucosal immune function. The aim of this study was to evaluate conjugated linoleic acid (CLA) on the Th17/Treg response and secretory IgA production in a model of giardiasis infection. Materials and Methods: C3H/HeN male mice were infected with 5×106 G. lamblia trophozoites (GS/M-83-H7, ATCC collection). Mice were assigned randomly to experimental and control groups. CLA was administered to the experimental group and phosphate-buffered saline (PBS) was given to the control group. Parasite load kinetics was determined. Enzyme-linked immunosorbent assay (ELISA) was performed to evaluate IgA and cytokines. Nuclear transcription factors and cytokines were measured by RT-qPCR, and histology of small bowel cells was evaluated. Results: CLA administration reduced the parasite load (P<0.05) and increased early Giardia-specific secretory IgA production. CLA also increased the expression of interleukin-10, transforming growth factor (TGF)-ß, and inducible nitric oxide synthase (iNOS) (P<0.05), while infection elevated the expression of Foxp3, with a peak at 40 days post-infection (P<0.05). There were no pathological changes in the colonic mucosa due to infection or treatment. Thus, CLA stimulated mucosal immunity and enhanced the humoral response against G. lamblia, not only for early infection control but also to promote regulatory cytokine production at 40 dpi, restoring the intestinal balance after parasite elimination. Conclusion: Our findings reveal novel anti-parasitic effects through the immune-modulatory activity of CLA against the intestinal parasite G. lamblia.

Influence of ferulic acid and clinoptilolite supplementation on growth performance, carcass, meat quality, and fatty acid profile of finished lambs.

This study was conducted to evaluate the effect of ferulic acid (FA) and clinoptilolite (CTL) supplementation on the growth performance, carcass characteristics, and meat quality of hair-breed lambs. Twenty-eight Kathadin male lambs (33.72 ± 3.4 kg) were randomly allocated to one of the four diets (n=7) under a 2 × 2 factorial arrangement to evaluate the effect of FA (0 or 300 ppm) and CTL (0% or 1%) during the last 40 days of the finishing phase. No interaction between additives was shown for growth performance, carcass characteristics and meat quality, with exception of the fatty acid profile (p < 0.05). FA reduced feed intake and carcass conformation (p < 0.05). Wholesale cuts were not affected by FA or CTL (p > 0.05). The L*, a*, and C* color parameters and some intramuscular fatty acids of the longissimus thoracis muscle were positively modified by CTL supplementation (p < 0.05). While there was no FA × CTL interaction, each additive could be used individually in animal nutrition to improve the feedlot performance and meat quality of the lambs.

ABCA1 gene R1587K polymorphism could be associated with metabolic syndrome and increased plasma triglyceride concentration in adults from northern Mexico / El polimorfismo R1587K del gen ABCA1 podría estar asociado con el síndrome metabólico y la concentración elevada de triglicéridos en adultos del norte de México

INTRODUCTION: the ABCA1 protein plays a key role in reverse cholesterol transport, promoting its clearance and high-density lipoprotein (HDL) biogenesis. The R1587K (rs2230808) single-nucleotide polymorphism (SNP) in the ABCA1 gene has been associated with dyslipidemia. OBJECTIVES: to investigate the relationship of R1587K genotypes with cardiovascular (CV) risk, metabolic syndrome (MetS), lipid profile, paraoxonase-1 (PON1) activity, and anti-oxLDL titers. METHODS: we performed a cross-sectional study in 57 northern Mexican adults with no reported diseases. The ABCA1 R1587K SNP was detected by real-time polymerase chain reaction (qPCR) using TaqMan allelic discrimination probes. We evaluated the relationship of R1587K with metabolic syndrome and clinical parameters including lipid profile, glucose and insulin, PON1 activity and concentration, anti-oxLDL antibodies, anthropometry and body-composition parameters, and the atherogenic index of plasma calculation. RESULTS: our results show higher triglyceride levels in the RK + KK carriers as compared to RR carriers (p = 0.031). An association between the RK + KK genotype and the presence of MetS (OR = 4.566, 95% CI = 1.386-14.92, p = 0.010) and a tendency towards high CV risk (OR = 3.317, 95% CI = 0.910-8.611, p = 0.069) was observed in comparison to RR carriers; however, there were no differences in HDL-C levels, PON1 activity and concentration, and anti-oxLDL titers among the R1587K genotypes. CONCLUSIONS: in the northern Mexican population, the ABCA1 gene R1587K SNP is present and the RK + KK genotypes are associated with MetS and increased triglyceride concentrations; therefore, it could be a CV risk biomarker. Nevertheless there is a need for further confirmation in longitudinal studies

INTRODUCCIÓN: la proteína ABCA1 juega un papel principal en el transporte reverso del colesterol, promoviendo su eliminación y la biogénesis de HDL. El polimorfismo de un solo nucleótido (SNP) R1587K (rs2230808) del gen ABCA1 se ha asociado con dislipidemias. OBJETIVO: investigar la relación de los genotipos del SNP R1587K con el riesgo cardiovascular (CV), el síndrome metabólico (SM), el perfil de lípidos, la actividad de paraoxonasa 1 (PON1) y los anticuerpos contra las LDL oxidadas (anti-oxLDL). MÉTODOS: se realizó un estudio transversal con 57 adultos del norte de México que reportaron no tener enfermedades diagnosticadas. El SNP R1587K del gen ABCA1 se detectó a través de PCR en tiempo real (qPCR) usando sondas TaqMan para discriminación alélica. Para evaluar la asociación del SNP R1587K con el SM y determinados parámetros clínicos se determinaron el perfil de lípidos, los niveles de glucosa e insulina, la actividad y concentración de PON1, los anticuerpos anti-oxLDL, los parámetros antropométricos y de composición corporal, y el cálculo del índice aterogénico en plasma. RESULTADOS: los resultados mostraron mayores niveles de triglicéridos en los portadores del genotipo RR + KK que en los portadores de RR (p = 0,031). Se observó una asociación entre el genotipo RK + KK y la presencia de SM (OR = 4,566, IC 95% = 1,386-14,92, p = 0,010) y una tendencia hacia un mayor riesgo cardiovascular (OR = 3,317, IC 95% = 0,910-8,611, p = 0,069) al compararlos con los portadores de RR. No se encontraron diferencias en los niveles de HDL-C, la actividad y concentración de PON1 y los anti-oxLDL entre los genotipos R1587K. CONCLUSIONES: el SNP R1587K del gen ABCA1 se encuentra presente en la población del norte de México y el genotipo RK + KK se asocia con el SM y concentraciones elevadas de triglicéridos, por lo que este SNP podría ser un biomarcador de riesgo cardiovascular. Sin embargo, se necesita confirmación a través de estudios longitudinales

ABCA1 gene R1587K polymorphism could be associated with metabolic syndrome and increased plasma triglyceride concentration in adults from northern Mexico.

INTRODUCTION: Objectives: to investigate the relationship of R1587K genotypes with cardiovascular (CV) risk, metabolic syndrome (MetS), lipid profile, paraoxonase-1 (PON1) activity, and anti-oxLDL titers. Methods: we performed a cross-sectional study in 57 northern Mexican adults with no reported diseases. The ABCA1 R1587K SNP was detected by real-time polymerase chain reaction (qPCR) using TaqMan allelic discrimination probes. We evaluated the relationship of R1587K with metabolic syndrome and clinical parameters including lipid profile, glucose and insulin, PON1 activity and concentration, anti-oxLDL antibodies, anthropometry and body-composition parameters, and the atherogenic index of plasma calculation. Results: our results show higher triglyceride levels in the RK + KK carriers as compared to RR carriers (p = 0.031). An association between the RK + KK genotype and the presence of MetS (OR = 4.566, 95% CI = 1.386-14.92, p = 0.010) and a tendency towards high CV risk (OR = 3.317, 95% CI = 0.910-8.611, p = 0.069) was observed in comparison to RR carriers; however, there were no differences in HDL-C levels, PON1 activity and concentration, and anti-oxLDL titers among the R1587K genotypes. Conclusions: in the northern Mexican population, the ABCA1 gene R1587K SNP is present and the RK + KK genotypes are associated with MetS and increased triglyceride concentrations; therefore, it could be a CV risk biomarker. Nevertheless there is a need for further confirmation in longitudinal studies.

INTRODUCCIÓN: Objetivo: investigar la relación de los genotipos del SNP R1587K con el riesgo cardiovascular (CV), el síndrome metabólico (SM), el perfil de lípidos, la actividad de paraoxonasa 1 (PON1) y los anticuerpos contra las LDL oxidadas (anti-oxLDL). Métodos: se realizó un estudio transversal con 57 adultos del norte de México que reportaron no tener enfermedades diagnosticadas. El SNP R1587K del gen ABCA1 se detectó a través de PCR en tiempo real (qPCR) usando sondas TaqMan para discriminación alélica. Para evaluar la asociación del SNP R1587K con el SM y determinados parámetros clínicos se determinaron el perfil de lípidos, los niveles de glucosa e insulina, la actividad y concentración de PON1, los anticuerpos anti-oxLDL, los parámetros antropométricos y de composición corporal, y el cálculo del índice aterogénico en plasma. Resultados: los resultados mostraron mayores niveles de triglicéridos en los portadores del genotipo RR + KK que en los portadores de RR (p = 0,031). Se observó una asociación entre el genotipo RK + KK y la presencia de SM (OR = 4,566, IC 95% = 1,386-14,92, p = 0,010) y una tendencia hacia un mayor riesgo cardiovascular (OR = 3,317, IC 95% = 0,910-8,611, p = 0,069) al compararlos con los portadores de RR. No se encontraron diferencias en los niveles de HDL-C, la actividad y concentración de PON1 y los anti-oxLDL entre los genotipos R1587K. Conclusiones: el SNP R1587K del gen ABCA1 se encuentra presente en la población del norte de México y el genotipo RK + KK se asocia con el SM y concentraciones elevadas de triglicéridos, por lo que este SNP podría ser un biomarcador de riesgo cardiovascular. Sin embargo, se necesita confirmación a través de estudios longitudinales. .

Hot environment on reproductive performance, immunoglobulins, vitamin E, and vitamin A status in sows and their progeny under commercial husbandry.

Hot environments can affect feed intake and lactation, and the subsequent unavailability of important micronutrients to the newborn piglet can impair piglet growth, reduce the viability of newborn piglets and limit their subsequent performance. This work addresses the effects of hot environments (summer season) upon the reproductive performance of sows during gestation and lactation as well as on the serum levels of vitamins and the concentration of immunoglobulins in their litters in comparison with the winter season. Fourteen sows were evaluated over 100 ± 2 days of gestation in each season. The temperature and humidity index (THI) was used as an indirect measure of heat stress during gestation. The reproductive performance, milk yield, and body condition of the sows were recorded. The concentrations of vitamin E and vitamin A in piglets and in sow serum, colostrum, milk and feed were determined by HPLC; immunoglobulins were assessed by an ELISA. The THI index indicated that animals were subject to heat stress only in during the summer. Although the effect was not significant, there were a lower number of piglets at birth and at weaning and the milk yield in summer compared with winter. There was no difference (p > 0.05) in the body condition of sows between seasons. Season had an effect (p < 0.05) on the vitamin A concentration of postpartum sow serum (0.29 µg/mL in winter vs 0.21 µg/mL in summer) and on the vitamin E concentration before birth (2.00 µg/mL in winter vs 0.90 µg/mL in summer). Vitamin E in milk was higher (p < 0.05) in winter than in summer (2.23 vs 1.81 µg/mL). Serum levels of vitamins A and E in piglets at birth were lower (p < 0.05) in winter than in summer. The concentrations of immunoglobulins (IgG and IgA) in colostrum and milk were similar between seasons (p > 0.05), but the IgA in piglet serum was higher in winter than in summer (p < 0.05). High temperatures produced heat stress in sows, which affected certain aspects of production that can be translated into economic losses for this sector.

Effect of gamma irradiation doses in the structural and functional properties of mice splenic cells.

PURPOSE: Ionizing radiation is nowadays effectively used in cancer treatments. However, the effect of irradiation in immune-system cells is poorly understood and remains controversial. The aim of this work was to determine the effect of γ-irradiation in the structural and functional properties of mice splenic cells. MATERIALS AND METHODS: Structural traits of irradiated splenic cells were evaluated by Atomic Force Microscopy and Raman spectroscopy. Functional properties were measured by gene and protein expression by RT-qPCR and ELISA, respectively. The induced cytotoxic effect was evaluated by MTT assay and the phagocytic capability by flow cytometry. RESULTS: Membrane roughness and molecular composition of splenic adherent cells are not changed by irradiation doses exposure. An increase in transcription of pro-inflammatory cytokines was observed. While protein expression decreased in IL-2 dose-dependent, relevant differences were identified in the anti-inflammatory marker IL-10 at 27 Gy. An increase of cytotoxicity in irradiated cells at 7 Gy and 27 Gy doses was observed, while phagocytosis was slight increased at 7 Gy dose but not statistically significant. CONCLUSIONS: We have demonstrated that γ-irradiation affects the splenic cells and changes the cytokines profile toward a pro-inflammatory phenotype and a tendency to increase the cytotoxicity was found, which implies a stimulation of immune response induced by γ-irradiation.

Central obesity and body fat, but not body mass index, are associated with the Pro12Ala polymorphism in the peroxisome proliferator-activated receptor γ gene in a population with a high consumption of saturated and trans-fatty acids.

The relationship of the Pro12Ala polymorphism in the peroxisome proliferator-activated receptor γ (PPARγ) gene with obesity and its modulation by dietary fat has been proposed, but the few studies addressing this issue have yielded controversial results. In a Mexican population characterized by high-fat consumption, we hypothesized that the Pro12Ala PPARγ genotype is related to obesity and this relationship is modulated by intake of saturated fatty acids (SFAs) and trans-fatty acids (TFAs). We recruited 69 adults for this cross-sectional study. The Pro12Ala PPARγ polymorphism was determined from blood genomic DNA by using real-time polymerase chain reaction. Univariate and multivariate logistic regression analyses were performed. Pro12Ala showed a positive association with central obesity (adjusted odds ratio, 7.38; 95% confidence interval, 1.19-45.77; P = .032) and percentage of body fat (%BF; adjusted odds ratio, 1.08; 95% confidence interval, 1.00-1.17; P = .048), suggesting that Pro12Ala carriers are more likely to have central obesity and a higher %BF than Pro12Pro carriers. A modifying effect was observed for the SFAs strata: we found a significant association between the Pro12Ala polymorphism and %BF in the high-SFA-intake stratum (P < .04), but not in the low-intake stratum (P > .7). No modifying effect was observed for the TFAs strata. In addition, the impact of total energy intake on obesity in Pro12Ala carriers seemed to be stronger than that in the wild-type genotype carriers. As hypothesized, our data demonstrated a relationship between the Pro12Ala PPARγ polymorphism and the presence of obesity, which is modulated by SFA intake but not TFA intake.

Serological and molecular prevalence of swine influenza virus on farms in northwestern Mexico.

The aim of this study was to provide an overview of the epidemiological status of swine influenza viruses in pigs from northwestern Mexico in 2008-2009. A serological and molecular survey was conducted in 150 pigs from 15 commercial farms in Sonora, Mexico (northwestern region of Mexico). The serological data showed that 55% of the sera were positive for the H1N1 subtype, 59% for the H3N2 subtype, and 38% for both subtypes. Overall, 16.6% (25/150) of the samples were positive for type A influenza by qRT-PCR. The phylogenetic analysis of the H1 viruses circulating in northwestern Mexico were grouped into cluster α, from five other clusters previously described. The influenza virus H1 circulating in northwestern Mexico showed 97-100% identity at the nucleotide level among them, 89% identity with other North American strains, 88% with strains from central Mexico, and 85% with the pandemic A/H1N1p2009 virus. Meanwhile, a closer relationship with some influenza viruses from North America (97% nucleotide identity) was found for H3 subtype. In conclusion, our results demonstrated a high circulation of strains similar to those observed in the North American linage among commercial farms in northwestern Mexico, involving of a different lineage virus different to the influenza pandemic of 2009.

Production of interferon α and ß, pro-inflammatory cytokines and the expression of suppressor of cytokine signaling (SOCS) in obese subjects infected with influenza A/H1N1.

BACKGROUND & AIMS: Obesity was recognized as an independent risk factor for morbidity and mortality during last influenza A/H1N1 pandemic. Mechanisms involved in the high mortality risk from obesity during influenza A virus include reduced type I interferon production and delayed pro-inflammatory response, which lead to a higher rate of morbidity and mortality in murine models. In this study, we evaluated the production of type I interferons, pro-inflammatory and anti-inflammatory cytokines in peripheral blood mononuclear cells (PBMCs) from obese and lean subjects with and without confirmed infection of influenza A/H1N1. The expression levels of the suppressor of cytokine signaling-1 (SOCS1), SOCS3 and nuclear factor-kB were also evaluated. METHODS: Cytokines were measured by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) and/or by ELISA in PBMCs stimulated with toll like receptor-3 (TLR-3) and TLR-7 ligands. The mRNA expression of SOCS1 and SOCS3 were evaluated by qRT-PCR. RESULTS: The obese volunteers infected with influenza A/H1N1 showed a diminished ability to produce type I interferon in response to TLR-3 ligand. Interestingly, the pro-inflammatory response was also affected in TLR-3 stimulated PBMCs. Obese influenza-free volunteers showed an increased basal expression of SOCS3, but not SOCS1. During influenza infection, SOCS1 and SOCS3 expression was higher in the lean infected volunteers in contrast to those who were obese infected. CONCLUSIONS: These data suggest that obesity is related to TLR-3 impairment and explain, at least in part, the inadequate immune response of obese individuals during infection with influenza A/H1N1 virus.

Decreased interferon-α and interferon-ß production in obesity and expression of suppressor of cytokine signaling.

OBJECTIVE: We analyzed the interferon-α (IFN-α), IFN-ß, and proinflammatory responses induced by Toll-like receptor (TLR) ligands in peripheral blood mononuclear cells (PBMCs) from obese subjects and their association with suppressor of cytokine signaling-1 (SOCS1) and SOCS3 expression. METHODS: The IFN responses were measured by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay in PBMCs stimulated with TLR-3 and TLR-7 ligands from 30 non-obese (body mass index ≤ 25 kg/m(2)) and 30 obese (body mass index ≥ 30 kg/m(2)) volunteers. The mRNA expression of nuclear factor-κB, SOCS1, and SOCS3 also was evaluated by qRT-PCR. Proinflammatory cytokine responses were measured by enzyme-linked immunosorbent assay. RESULTS: Obese subjects showed a decreased ability to produce IFN-α and IFN-ß in response to TLR ligands; this response was associated with increased basal levels of SOCS3 but not SOCS1. However, after stimulation, the expression of SOCS3 and SOCS1 mRNA was significantly lower in PBMCs from obese compared with non-obese subjects. The PBMCs from obese subjects also showed higher basal levels of interleukin-6 and a decreased response of proinflammatory cytokines interleukin-6 and interleukin-1ß after stimulation with the TLR-3 ligand compared with PBMCs from non-obese participants. CONCLUSION: These data suggest that obesity is related to impaired IFN-α and IFN-ß responses and increased SOCS3 basal mRNA expression and that a signaling pathway by TLR-3 may be involved. These results could explain, at least in part, the inadequate response of obese people against viral infections, such as influenza.

Expresión de citocinas en cerdos co-infectados con circovirus porcino tipo 2 y el virus del síndrome respiratorio y reproductivo porcino / Cytokine expression in growing pigs co-infected with porcine circovirus type 2 and porcine reproductive and respiratory syndrome virus

The aim of this work was to quantify porcine circovirus type 2 (PCV2), porcine reproductive and respiratory syndrome virus (PRRSV), as well as cytokine mRNA expression (IL-6, IL-12, TNF-α, and IFN-γ) in superficial inguinal lymph nodes (SILN) of pigs from postweaning multisystemic wasting syndrome (PMWS)-affected and healthy farms. Genetic characterization of detected PCV2 sequences was also carried out. Based on the clinical outcome and the quantification of PCV2 nucleic acid by in situ hybridization and real time PCR, pigs were grouped into three categories: 1) PMWS, pigs with signs of wasting and high viral load in SILN (n = 4); 2) wasted-non-PMWS, pigs with signs of wasting and low to intermediate viral loads in SILN (n = 3); and 3) healthy, pigs with no clinical signs and low viral loads (n = 3). PRRSV was detected in three PMWS affected and two-wasted-non-PMWS pigs. The genetic analysis of PCV2 sequences revealed the presence of two genotypes PCV2a and PCV2b in PMWS-affected farms. Cytokine mRNA expression revealed that PMWS affected pigs had low expression of IFN-γ, whereas wasted-non-PMWS pigs showed higher amounts of IFN-γ. These results suggest that an imbalance in cytokines could be involved in the pathogenesis of PMWS.

Los objetivos de este trabajo fueron la cuantificación de circovirus porcino tipo 2 (PCV2) de virus del síndrome respiratorio y reproductivo porcino (PRRSV), así como la expresión de citocinas (IL-6, IL-12, TNF-α, e IFN-γ) en nódulos linfáticos inguinales superficiales (NLIS) de cerdos afectados por el síndrome de desmedro posdestete (postweaning multisystemic wasting syndrome, PMWS) y en cerdos sanos. Con base en el diagnóstico clínico y a la cuantificación de PCV2 por hibridación in situ y qRT-PCR, los cerdos se distribuyeron en tres grupos: 1) PMWS, cerdos con signos de desmedro y con cargas virales altas en NLIS (n = 4); 2) con desmedro sin PMWS, cerdos con signos de desmedro y cargas virales de intermedias a bajas en NLIS (n = 3); y 3) cerdos sanos, sin signos clínicos y con cargas virales bajas (n = 3). El PRRSV fue detectado en tres de los cerdos afectados por el PMWS y en dos cerdos con desmedro, pero sin el síndrome. Se caracterizaron las secuencias nucleotídicas del ORF2 de los PCV2 encontrados y los análisis genéticos revelaron la presencia de 2 genotipos PCV2a y PCV2b en las granjas afectadas con el PMWS. El perfil de expresión de citocinas mostró una baja expresión de IFN-γ en los cerdos con PMWS, mientras que los cerdos con desmedro sin PMWS mostraron valores elevados de esta citocina. Estos resultados sugieren que existe un desbalance en la producción de citocinas que puede estar implicado en la patogénesis de la enfermedad.

Genetic analysis of avian influenza virus from migratory waterfowl in Mexico.

This study describes the genetic characterization of avian influenza virus from waterfowl in Mexico. Partial sequences of two H5, one H6, and one H9 hemmaglutinin (HA) genes were determined. The deduced amino acid sequences showed that they were low-pathogenic viruses (LPAI). Phylogenetic analysis of H5 and H6 HA indicates a North American lineage, closely related to contemporary Californian isolates, and H9 HA was closer to the Korean-like lineage. These results demonstrate the introduction of a diverse genetic pool of subtypes to Sonora estuaries through circulation of bird species carriers from the Pacific flyway.

Estandarización de un método de detección molecular del virus influenza (H5N1) de alta patogenicidad / Standardization of a molecular detection method of highly pathogenic avian influenza virus (H5N1)

El virus de la influenza aviar H5N1 de alta patogenicidad mantiene el alerta mundial debido a su potencial zoonótico y pandémico. Surge entonces la necesidad de contar con herramientas para la detección temprana y de esta forma reducir el impacto potencial a la salud humana y animal. En este estudio se estandarizó un método de detección molecular de los genes de la matriz (M), hemaglutinina (H5) y neuraminidasa (N1) del virus de la influenza aviar H5N1 de alta patogenicidad de linaje asiático, mediante transcripción-reversa y reacción en cadena de la polimerasa en tiempo real (RRT-PCR). A partir de un ARN viral de referencia cepa A/Vietnam/1203/2004 (H5N1) se construyeron controles positivos mediante clonación de productos de PCR. Los estándares de naturaleza plasmídica se emplearon en la obtención de curvas estándar para determinar los límites de detección de la técnica. La sensibilidad observada para todos los genes analizados fue de 10² copias de ADN/μL. Las curvas mostraron una eficiencia superior al 90%, y R²>0,99. Este método puede ser útil en las campañas de monitoreo del virus en aves migratorias, así como para el tamizaje de muestras clínicas de humanos, en una emergencia de salud.

Highly pathogenic avian influenza virus (HPAI) H5N1 is a global threat due to its zoonotic and pandemic potential. Then, concern arises and the need to have early detection tools to minimize the impact on human and animal health. In this work, a molecular detection method was implemented to detect matrix (M), hemagglutinin (H5) and neuraminidase (N1) genes of HPAI avian influenza virus H5N1, based on real time RT-PCR (RRT-PCR). Positive controls were constructed from reference RNA viral A/Vietnam/1203/2004 (H5N1), cloned into plasmidic vectors and sequenced. Assay detection sensitivity was assessed with standard curves for each gene. Assay sensitivity was 10² DNA copies/μl in all cases. Curves showed amplification efficiency higher than 90% and R²>0.99. This method could be useful for bird monitoring campaigns and as a screening procedure for clinical samples.