A Proteinaceous Alpha-Amylase Inhibitor from Moringa Oleifera Leaf Extract: Purification, Characterization, and Insecticide Effects against C. maculates Insect Larvae.

The main objective of the current study was the extraction, purification, and enzymatic characterization of a potent proteinaceous amylase inhibitor from Moringa oleifera. The antimicrobial potential and insecticide effects against C. maculates insect larvae were also studied. The α-amylase inhibitor was extracted in methanol (with an inhibitory activity of 65.6% ± 4.93). Afterwards, the inhibitor αAI.Mol was purified after a heat treatment at 70 °C for 15 min followed by one chromatographic step of Sephadex G-50. An apparent molecular weight of 25 kDa was analyzed, and the N-terminal sequence showed the highest identity level (89%) with the monomeric α-amylase inhibitor from Triticum dicoccoides. αAI.Mol was found to tolerate pH values ranging from 5.0 to 11.0 and showed maximal activity at pH 9.0. Thermal stability was remarkably important, since the inhibitory activity was maintained at 55% after 1 h of incubation at 70 °C and at 53% after an incubation of 45 min at 80 °C. The potency of the current purified inhibitor against amylases from different origins indicates that αAI.Mol seems to possess the highest affinity toward human salivary α-amylase (90% inhibitory activity), followed by the α-amylase of insects Callosobruchus maculatus and Tribolium confusum (71% and 61%, respectively). The kinetic parameters were also calculated, and the Kmax and Vmax of the digestive amylase were estimated at 185 (mmol/min/mg) and 0.13 mM, respectively. The inhibitor possesses a strong bactericidal effect against Gram+ and Gram- strains, and the MIC values were >1 against B. cereus but >6 against E. coli. Interestingly, the rates of survival and pupation of C. maculates insect larvae were remarkably affected by the purified αAI.Mol from Moringa oleifera.

Biochemical, Kinetic and Biological Properties of Group V Phospholipase A2 from Dromedary.

Secretory group V phospholipase A2 (PLA2-V) is known to be involved in inflammatory processes in cellular studies, nevertheless, the biochemical and the enzymatic characteristics of this important enzyme have been unclear yet. We reported, as a first step towards understanding the biochemical properties, catalytic characteristics, antimicrobial and cytotoxic effects of this PLA2, the production of PLA2-V from dromedary. The obtained DrPLA2-V has an absolute requirement for Ca2+ and NaTDC for enzymatic activity with an optimum pH of 9 and temperature of 45 °C with phosphatidylethanolamine as a substrate. Kinetic parameters showed that Kcat/Kmapp is 2.6 ± 0.02 mM-1 s-1. The enzyme was found to display potent Gram-positive bactericidal activity (with IC50 values of about 5 µg/mL) and antifungal activity (with IC50 values of about 25 µg/mL)in vitro. However, the purified enzyme did not display a cytotoxic effect against cancer cells.

Efficient heterologous expression in Pichia pastoris, immobilization and functional characterization of a scorpion venom secreted phospholipase A2.

Industrial processes have expanded with the ability to clone and express recombinant immobilized enzymes in microorganisms such as Pichia pastoris that have commercially attractive amounts of the appropriate genes. This report describes the overexpression in Pichia pastoris, immobilization, and functional characterization of a secreted phospholipase A2 from scorpion venom Scorpio maurus: rPLA2(-5). After 48 h of culture, the recombinant rPLA2(-5) was secreted into the culture medium and expressed at about 9 mg/L. Comparative analyses of the kinetics and hydrolysis of rPLA2(-5) monolayers at various surface pressures were conducted with the same form produced in Escherichia coli. As a second part of the study, rPLA2(-5) overexpressed in Pichia pastoris was immobilized by adsorption on CaCO3, with about 78 percent of the activity. In comparison to the free enzyme, rPLA2(-5) was studied for stability. Immobilization improved the thermal stability of rPLA2(-5) and even the stability at acidic pH. Moreover, we found that the immobilization improved the stability of rPLA2(-5) towards bile salts, Tween 80, Triton X-100, and SDS, as well as its stability towards many organic solvents. Until now, this is the first study to describe the overexpression and immobilization of a scorpion venom phospholipase A2 that possesses an interesting stability characteristic that makes it useful for a wide range of biotechnological applications.

Production and structure prediction of amylases from Chlorella vulgaris.

Amylases are enzymes required for starch degradation and are naturally produced by many microorganisms. These enzymes are used in several fields such as food processing, beverage, and medicine as well as in the formulation of enzymatic detergents proving their significance in modern biotechnology. In this study, a three-stage growth mode was applied to enhance starch production and amylase detection from Chlorella vulgaris. Stress conditions applied in the second stage of cultivation led to an accumulation of proteins (75% DW) and starch (21% DW) and a decrease in biomass. Amylase activities were detected and they showed high production levels especially on day 3 (35 U/ml) and day 5 (22.5 U/ml) of the second and third stages, respectively. The bioinformatic tools used to seek amylase protein sequences from TSA database of C. vulgaris revealed 7 putative genes encoding for 4 α-amylases, 2 ß-amylases, and 1 isoamylase. An in silico investigation showed that these proteins are different in their lengths as well as in their cellular localizations and oligomeric states though they share common features like CSRs of GH13 family or active site of GH14 family. In brief, this study allowed for the production and in silico characterization of amylases from C. vulgaris.

A Novel Thermostable and Alkaline Protease Produced from Bacillus stearothermophilus Isolated from Olive Oil Mill Sols Suitable to Industrial Biotechnology.

This study was conducted to identify a new alkaline and thermophilic protease (Ba.St.Pr) produced from Bacillus stearothermophilus isolated from olive oil mill sols and to evaluate its culture conditions, including temperature, pH, carbon and nitrogen sources, and incubation time. The optimum culture conditions for cell growth (10 g/L) and protease production (5050 U/mL) were as follows: temperature 55 °C, pH 10, inoculation density 8 × 108 CFU/mL, and incubation time 24 h. The use of 3% yeast extract as the nitrogen sources and galactose (7.5 g/L) as the carbon sources enhanced both cell growth and protease production. Using reversed-phase analytical HPLC on C-8 column, the new protease was purified with a molecular mass of approximately 28 kDa. The N-terminal sequence of Ba.St.Pr exhibited a high level of identity of approximately 95% with those of Bacillus strains. Characterization under extreme conditions revealed a novel thermostable and alkaline protease with a half-life time of 187 min when incubated with combined Ca2+/mannitol. Ba.St.Pr demonstrated a higher stability in the presence of surfactant, solvent, and Ca2+ ions. Consequently, all the evaluated activity parameters highlighted the promising properties of this bacterium for industrial and biotechnological applications.

Molecular and Structural Characterizations of Lipases from Chlorella by Functional Genomics.

Microalgae have been poorly investigated for new-lipolytic enzymes of biotechnological interest. In silico study combining analysis of sequences homologies and bioinformatic tools allowed the identification and preliminary characterization of 14 putative lipases expressed by Chlorella vulagaris. These proteins have different molecular weights, subcellular localizations, low instability index range and at least 40% of sequence identity with other microalgal lipases. Sequence comparison indicated that the catalytic triad corresponded to residues Ser, Asp and His, with the nucleophilic residue Ser positioned within the consensus GXSXG pentapeptide. 3D models were generated using different approaches and templates and demonstrated that these putative enzymes share a similar core with common α/ß hydrolases fold belonging to family 3 lipases and class GX. Six lipases were predicted to have a transmembrane domain and a lysosomal acid lipase was identified. A similar mammalian enzyme plays an important role in breaking down cholesteryl esters and triglycerides and its deficiency causes serious digestive problems in human. More structural insight would provide important information on the enzyme characteristics.

Spirulina platensis, Punica granatum peel, and moringa leaves extracts in cosmetic formulations: an integrated approach of in vitro biological activities and acceptability studies.

The selection of suitable natural raw materials in the cosmetic research and development is a key point, in order not only to obtain the expected results but also to avoid undesirable side effects. In this study, spirulina platensis, pomegranate (Punica granatum) peel, and moringa leaves alone were evaluated for anti-oxidant and antimicrobial properties. The chemical composition (moisture, dry matter, protein, lipid, and ash) and total polyphenols, flavonoids, and carotenoids content were evaluated in the three extracts. Total antioxidant capacity and ferric reducing activity power of extracts were also studied. Using agar diffusion method, the anti-Micrococcus luteus, Staphylococcus aureus, E. coli, Listeria monocytogenes, Salmonella typhimurium, and Enterococus faecalis activities were measured. Interestingly, after combinations, pomegranate peel/spirulina (A), and moringa/spirulina (B): 25%/75% and 50%/50%, we have found that pomegranate peel can be incorporated into cosmetic formulations as an excellent preservative due to its exceptionally amount of phenolic compounds, powerful antioxidant activity, and its antibacterial activity against pathogenic strains.

Purification and Biochemical Characterization of a New Protease Inhibitor from Conyza dioscoridis with Antimicrobial, Antifungal and Cytotoxic Effects.

The main objective of the current study was the extraction, purification, and biochemical characterization of a protein protease inhibitor from Conyzadioscoridis. Antimicrobial potential and cytotoxic effects were also examined. The protease inhibitor was extracted in 0.1 M phosphate buffer (pH 6-7). Then, the protease inhibitor, named PDInhibitor, was purified using ammonium sulfate precipitation followed by filtration through a Sephadex G-50 column and had an apparent molecular weight of 25 kDa. The N-terminal sequence of PDInhibitor showed a high level of identity with those of the Kunitz family. PDInhibitor was found to be active at pH values ranging from 5.0 to 11.0, with maximal activity at pH 9.0. It was also fully active at 50 °C and maintained 90% of its stability at over 55 °C. The thermostability of the PDInhibitor was clearly enhanced by CaCl2 and sorbitol, whereas the presence of Ca2+ and Zn2+ ions, Sodium taurodeoxycholate (NaTDC), Sodium dodecyl sulfate (SDS), Dithiothreitol (DTT), and ß-ME dramatically improved the inhibitory activity. A remarkable affinity of the protease inhibitor with available important therapeutic proteases (elastase and trypsin) was observed. PDInhibitor also acted as a potent inhibitor of commercial proteases from Aspergillus oryzae and of Proteinase K. The inhibitor displayed potent antimicrobial activity against gram+ and gram- bacteria and against fungal strains. Interestingly, PDInhibitor affected several human cancer cell lines, namely HCT-116, MDA-MB-231, and Lovo. Thus, it can be considered a potentially powerful therapeutic agent.

Enhancement of Aeribacillus pallidus strain VP3 lipase catalytic activity through optimization of medium composition using Box-Behnken design and its application in detergent formulations.

Lipases are hydrolytic enzymes owing much importance in industrial applications. These enzyme-based detergents are ecofriendly and produce a wastewater with low level of COD (chemical oxygen demand). In the present work, a novel halophilous, thermoalkaline, and detergent-tolerant lipase produced by a newly isolated Aeribacillus pallidus strain VP3 was studied. Considerable interest has been given to this lipase by the improvement of its catalytic activity through the optimization of the pH, the (C/N) ratio, and the inoculum size, using the response surface methodology based on the Box-Behnken design of experiments. A total of 16 experiments were conducted, and the optimized pH, (C/N) ratio, and inoculum size were 10, 1, and 0.3, respectively. The results of the analysis of variance (ANOVA) test indicated that the established model was significant (p value < 0.05). The optimization of the production conditions leads to 2.83-fold of increase in the catalytic activity calculated as the ratio of the activity obtained after optimization (68 U) and the initial activity before optimization (24 U). All in all, the lipase of Aeribacillus pallidus could be considered as a potential candidate to be incorporated in detergent formulations since it shows a good stability towards detergents and wash performance.

Hydrolysis of three different head groups phospholipids by chicken group V phospholipase A2 using the monomolecular film technique.

The kinetic aspects of lipolysis by pulmonary phospholipase A2 (ChPLA2-V), chicken intestinal phospholipase A2 (ChPLA2-IIA) and chicken pancreatic phospholipase A2 (ChPLA2-IB), from chicken have been compared using the monomolecular films technique, on short-chain phospholipids (with three different head groups) and on long-chain phospholipids. The main conclusions from our experimental data indicate that the maximum catalytic activities of ChPLA2-V on 1,2 phosphatidylcholine and 1,2 phosphatidylethanolamine reached 15.26 and 36.12 moles/cm2.min.mM, respectively, at a pressure of 15 and 35 dynes/cm, respectively. Whereas, those of ChPLA2-IB were 3.58 (at the pressure of 20 dynes/cm) and 4.9 moles/cm2.min.mM. However, hydrolysis of phosphatidylglycerol monolayers (C12PG), were very much higher compared with all the substrates tested with 122 moles/cm2.min. Surprisingly, the hydrolysis rate of ChPLA2-V on long-chain phosphatidylglycerol (C18PG) was very low (1.45 moles/cm2.min) compared with all tested substrates, even with the use of p-cyclodextrin. And thus, the fatty acid preference of ChPLA2-V was 2-decanoyl > 2-oleoyl with a PG head group. In order to gain significant correlations between enzyme's structures and their relative functions, we tried to examine the surface electrostatic potentials of the various secreted phospholipase 2 (sPLA2) from chicken. In the present study, we detailed that the substrate affinity, specificity and the hydrolysis rates of sPLA2 at each interface is governed by the surface electrostatic potentials and hydrophobic interactions operative at this surface.

Purification, biochemical and molecular study of lipase producing from a newly thermoalkaliphilic Aeribacillus pallidus for oily wastewater treatment.

Treatment of oily wastewater is constantly a challenge; biological wastewater treatment is an effective, cheap and eco-friendly technology. A newly thermostable, haloalkaline, solvent tolerant and non-induced lipase from Aeribacillus pallidus designated as GPL was purified and characterized of biochemical and molecular study for apply in wastewater treatment. The GPL showed a maximum activity at 65°C and pH 10 after 22 h of incubation, with preference to TC4 substrates. Pure enzyme was picked up after one chromatographic step. It displayed an important resistance at high temperature, pH, NaCl, at the presence of detergents and organic solvents. In fact, GPL exhibited a prominent stability in wide range of organic solvents at 50% (v/v) concentration for 2 h of incubation. The efficiency of the GPL in oil wastewater hydrolysis was established at 50°C for 1 h, the oil removal efficiency was established at 96, 11% and the oil biodegradation was confirmed through fourier transform infrared (FT-IR) spectroscopy. The gene that codes for this lipase was cloned and sequenced and its open reading frame encoded 236 amino acid residues. The deduced amino acids sequence of the GPL shows an important level of identity with Geobacillus lipases.

A novel bactericidal homodimeric PLA2 group-I from Walterinnesia aegyptia venom.

A novel non-toxic phospholipase A2 was purified to homogeneity in a single chromatography step from the venom of Walterinnesia aegyptia, a monotypic elapid snake caught in Saudi Arabia, and its antimicrobial and hemolytic properties were evaluated as well. This enzyme, namely WaPLA2, is a homodimer with an estimated molecular mass of 30 kDa, and its NH2-terminal sequence exhibits a significant degree of similarity with PLA2 group-I. At optimal pH (8.5) and temperature (45 °C), the purified PLA2 exhibited a specific activity of 2100 U/mg, and it requires bile salts and Ca2+ for its activity. However, other cations such as Cd2+ and Hg2+ diminished the enzyme activity remarkably, thereby suggesting that the catalytic site arrangement has an exclusive structure for Ca2+ binding. Furthermore, WaPLA2 maintained almost 100% and 60% of its full activity in a pH range of 6.0-10 after 24 h incubation or after 60 min treatment at 70 °C, respectively. In the biological activity assays, WaPLA2 displayed potent indirectly hemolytic and antimicrobial activities that were strongly correlated. These promising findings encourage further in-depth research to understand the molecular mechanism of WaPLA2's antimicrobial properties for its possible use as a potential therapeutic lead molecule for treating infections.

Antibacterial, antifungal and anticoagulant activities of chicken PLA2 group V expressed in Pichia pastoris.

Secretory class V phospholipase A2 (PLA2-V) has been shown to be involved in inflammatory processes in cellular studies, but the biochemical and physical properties of this important enzyme have been unclear. As a first step towards understanding the structure, function and regulation of this PLA2, we report the expression and characterization of PLA2-V from chicken (ChPLA2-V). The ChPLA2-V cDNA was synthesized from chicken heart polyA mRNA by RT-PCR, and an expression construct containing the PLA2 was established. After expression in Pichia pastoris cells, the active enzyme was purified. The purified ChPLA2-V protein was biochemically and physiologically characterized. The recombinant ChPLA2-V has an absolute requirement for Ca2+ for enzymatic activity. The optimum pH for this enzyme is pH 8.5 in Tris-HCl buffer with phosphatidylcholine as substrate. ChPLA2-V was found to display potent Gram-positive and Gram-negative bactericidal activity and antifungal activity in vitro. The purified enzyme ChPLA2-V with much stronger anticoagulant activity compared with the intestinal and pancreatic chicken PLA2-V was approximately 10 times more active. Chicken group V PLA2, like mammal one, may be considered as a future therapeutic agents against fungal and bacterial infections and as an anticoagulant agent.

Maneb disturbs expression of superoxide dismutase and glutathione peroxidase, increases reactive oxygen species production, and induces genotoxicity in liver of adult mice.

Maneb (MB), a fungicide largely used in agriculture throughout the world including Tunisia, protects many vegetables, fruits and field crops against a wide spectrum of fungal diseases. However there is a lack of informations regarding the risks arising from MB exposure on non target organisms, especially mammals. The aim of this study was to investigate the morphological, biochemical and molecular aspects of liver injury after exposure of mice to MB. Four doses of MB corresponding to 1/8 (group D1), 1/6 (group D2), 1/4 (group D3), and 1/2 (group D4) of lethal dose (DL50 = 1500 mg/kg body weight) were administered to adult mice. Oxidative stress parameters were also objectified by molecular and histological endpoints in the liver. Maneb caused hepatotoxicity as characterized by the significant increase in the levels of malondialdehyde and protein oxidation marker, advanced oxidation protein products (AOPP). The activities of catalase, glutathione peroxidase, superoxide dismutase and the levels of glutathione decreased significantly in all treated mice, while vitamin C levels decreased only in group D4. We also noted a significant decrease in gene expression of superoxide dismutase and glutathione peroxidase enzymes. Maneb caused nucleic acids degradation testifying its genotoxicity. Yet, biochemical markers in plasma showed a decrease in total protein and an increase in aspartate, alanine amino transferases and bilirubin levels in all treatment groups. Moreover, plasma levels of cholesterol, triglycerides and low density lipoprotein-cholesterol significantly increased, while those of high density lipoprotein-cholesterol decreased. These biochemical alterations were correlated with significantly histological changes. Our data showed, for the first time, that intraperitoneal injection of very high non environmentally relevant MB concentrations to adult mice resulted in oxidative stress leading to hepatotoxicity and the impairment of defense systems, confirming the pro-oxidant and genotoxic effects of this fungicide.

Renaturation and one step purification of the chicken GIIA secreted phospholipase A2 from inclusion bodies.

The cDNA coding for a mature protein of 123 amino acids, containing all of the structural features of catalytically active group IIA sPLA2, has been amplified from chicken intestine. The gene has been cloned into the bacterial expression vector pET-21a(+), which allows protein over-expression as inclusion bodies and enables about 3mg/l of pure refolded fully active enzyme to be obtained. Recombinant expression of chicken intestinal sPLA2-IIA (ChPLA2-IIA) in Escherichia coli shows that the enzyme is Ca(2+) dependent, maximally active at pH 8-9, and hydrolyses phosphatidylglycerol versus phosphatidylcholine with a 10-fold preference. Indeed, we report in this work, a comparative kinetic study between the wild type and the recombinant ChPLA2-IIA, on zwitterionic head group phospholipids (DDPC) and negatively charged phospholipids (POPG) using the monomolecular film technique. The ability to express reasonably large amounts of the sPLA2 Group IIA, compared to that obtained with the classical purification will provide a basis for future site directed mutagenesis studies of this important enzyme.

Dimethoate induces kidney dysfunction, disrupts membrane-bound ATPases and confers cytotoxicity through DNA damage. Protective effects of vitamin E and selenium.

Dimethoate (DM) is an organophosphate insecticide widely used in agriculture and industry and has toxic effects on non-target organisms especially mammalian. However, we still know little about DM-induced kidney injury and its alleviation by natural antioxidants. In the present study, selenium (Se), vitamin E, DM, Se+DM, vitamin E+DM, Se+vitamin E+DM were given to adult rats for 4 weeks. Plasma creatinine and uric acid, kidney MDA, PC, H2O2 and AOPP levels were higher, while Na(+)-K(+)-ATPase and LDH values were lower in the DM group than those of controls. A smear without ladder formation on agarose gel was shown in the DM group, indicating random DNA degradation and DM-induced genotoxicity. A decrease in kidney GSH, NPSH and plasma urea levels and an increase in GPx, SOD and catalase activities were observed in the DM group when compared to those of controls. Plasma cystatin C levels increased, indicating a decrease in glomerular filtration rate. When Se or vitamin E was added through diet, the biochemical parameters cited above were partially restored in Se+DM and vitamin E+DM than DM group. The joint effect of Se and vitamin E was more powerful against DM-induced oxidative stress and kidney dysfunction. The changes in biochemical parameters were substantiated by histological data. In conclusion, our results indicated a possible mechanism of DM-induced nephrotoxicity, where renal genotoxicity was noted, membrane-bound ATPases and plasma biomarkers were disturbed. Se and vitamin E ameliorated the toxic effects of this pesticide in renal tissue suggesting their role as potential antioxidants.

N-terminal domain of turkey pancreatic lipase is active on long chain triacylglycerols and stabilized by colipase.

The gene encoding the TPL N-terminal domain (N-TPL), fused with a His6-tag, was cloned and expressed in Pichia pastoris, under the control of the glyceraldehyde-3-phosphate dehydrogenase (GAP) constitutive promoter. The recombinant protein was successfully expressed and secreted with an expression level of 5 mg/l of culture medium after 2 days of culture. The N-TPL was purified through a one-step Ni-NTA affinity column with a purification factor of approximately 23-fold. The purified N-TPL, with a molecular mass of 35 kDa, had a specific activity of 70 U/mg on tributyrin. Surprisingly, this domain was able to hydrolyse long chain TG with a specific activity of 11 U/mg using olive oil as substrate. This result was confirmed by TLC analysis showing that the N-TPL was able to hydrolyse insoluble substrates as olive oil. N-TPL was unstable at temperatures over 37°C and lost 70% of its activity at acid pH, after 5 min of incubation. The N-TPL exhibited non linear kinetics, indicating its rapid denaturation at the tributyrin-water interface. Colipase increased the N-TPL stability at the lipid-water interface, so the TPL N-terminal domain probably formed functional interactions with colipase despite the absence of the C-terminal domain.

Renaturation and one step purification of the chicken GIIA secreted phospholipase A2 from inclusion bodies.

The cDNA coding for a mature protein of 123 amino acids, containing all of the structural features of catalytically active group II sPLA2, has been amplified. The gene has been cloned into the bacterial expression vector pET-21a(+), which allows protein over-expression as inclusion bodies and enables about 3 mg per litre of pure refolded fully active enzyme to be obtained. Recombinant expression of chPLA2-IIA in Escherichia coli shows that the enzyme is Ca(2+) dependent, maximally active at pH 8-9, and hydrolyses phosphatidylglycerol versus phosphatidylcholine with a 15-fold preference. The ability to express reasonably large amounts of the sPLA2 Group IIA, compared to that obtained with the classical purification will provide a basis for future site directed mutagenesis studies of this important enzyme.

Phospholipase A2 purification and characterization: a case study.

We compared here the purification procedures, the pH, the calcium, the bile salts, and the temperature dependencies as well as the catalytic activities on phosphatidylcholine (PC) and phosphatidylethanolamine (PE) of two purified secreted PLA2 from chicken pancreatic (ChPLA2-IB) and chicken intestinal (ChPLA2-IIA) origins. Interestingly, ChPLA2-IB hydrolyzes efficiently both purified PC and PE, whereas ChPLA2-IIA hydrolyzes only PE and not PC, even after a long incubation period. These analytical results clearly indicate that the catalytic activity of ChPLA2-IIA, measured with the pH-stat and using egg yolk as substrate, is mainly due to the hydrolysis of the PE fraction present in egg yolk.

Drastic changes in the tissue-specific expression of secreted phospholipases A2 in chicken pulmonary disease.

Infectious bronchitis is one of the most important diseases in poultry and it causes major economic losses. Infectious bronchitis is an acute, highly contagious, viral disease of chickens, characterized by rales, coughing, and sneezing. Because secreted phospholipases A2 (sPLA2) are involved in inflammatory processes, the gene expressions of sPLA2s were investigated in both healthy chickens and chickens with infectious bronchitis and lung inflammation. The draft chicken genome was first scanned using human sPLA2 sequences to identify chicken sPLA2s (ChPLA2), chicken total mRNA were isolated and RT-PCR experiments were performed to amplify and then sequence orthologous cDNAs. Full-length cDNA sequences of ChPLA2-IB, -IIA, -IIE, -V and -X were cloned. The high degree of sequence identity of 50-70% between the avian and mammalian (human and mouse) sPLA2 orthologs suggests a conservation of important enzymatic functions for these phospholipases. Quantitation by qPCR of the transcript levels of ChPLA2-IB, -IIA, -IIE, -V and -X in several tissues from healthy chicken indicated that the expression patterns and mRNA levels diverged among the phospholipases tested. In chicken with infectious bronchitis, an over expression of ChPLA2-V was observed in lungs and spleen in comparison with healthy chicken. These findings suggest that ChPLA2-V could be a potential biomarker for lung inflammation. Conversely, a down regulation of ChPLA2-IB, -IIA and -X was observed in lungs and spleen in case of infectious bronchitis. A significant increase in the expression level of ChPLA2-X and ChPLA2-IB was also noticed in pancreas. No or minor changes have been detected in the expression of ChPLA2-IIE in lungs and small intestine, but it shows a significant increase in several infected tissues.