Production of an Antibody Fragment (scFv) Targeting PcrV Protein of Pseudomonas aeruginosa in Fed-Batch Cultivation Mode

Background: Pseudomonas aeruginosa is one of the opportunistic pathogens causing frequent hospital-acquired life-threatening infections in mechanically ventilated patients. The most significant virulence factor of P. aeruginosa is T3SS. PcrV is an important structural protein of the T3SS. Methods:In the current investigation, a recombinant scFv mAb against the PcrV protein was expressed in EnBase® (fed-batch) cultivation mode. The pETiteTM N-His SUMO Kan vector, including anti-PcrV scFv gene, was transformed into Escherichia coli (BL21) cells. The expression and solubility of anti-PcrV scFv protein were investigated at two different temperatures (25 °C and 30 °C) and at different induction times (4, 6, 8, 12, and 24 hours). Results:Increased efficiency was achieved by EnBase® compared to LB broth; owing to the slow release of glucose, the maximum level of solubility and total protein expression was observed in EnBase® cultivation system at 30 °C and 24 h post induction. Furthermore, IC₅₀ for anti-PcrV scFv protein was determined to be approximately 7 μg/mL. Conclusion:Anti-PcrV scFv produced in this study showed promising in vitro results, protecting RBC from lysis by P. aeruginosa (exoU+). Key Words: Fed Batch, recombinant protein, Pseudomonas aeruginosa, scFv     

The study lentil adaptations to highland winter-sown environments in Northeastern Iran

One hundred lentil genotypes accessions of Mashhad Lentil Collection (MLC) were evaluated at Chenaran, Iran with three planting dates [10 Oct., 10 Nov. (fall) and 28 Mar. (spring) during 2008/09 growing seasons]. Evaluation based on winter survival percentage showed that in 2008/09 growing season, percentage of highly tolerant, tolerant, moderately tolerant, moderately susceptible and susceptible genotypes were 22, 61, 16, 1 and 0 respectively. In the first planting date more than 50% of genotypes were ranked as highly tolerant, however it was decreased to 12% and 5% of genotypes in the second and third planting dates, respectively, in this year. According to the fall plantings data, the length of growth duration of samples in fall plantings were 2.3 times higher than spring planting, and also in the first planting that was 5, 25 and 255 percent more than 2^nd and 3^rd planting dates, respectively. The height of fall plants were more than spring plants and in the first year fall planting genotypes were about 38% taller than the same planting. The number of pod per plant in fall planting was 2 times higher than spring planting. More than 22% of accessions had 125 pods per plant in 1^st and 2^nd planting in this year. In the 2008/09 growing seasons the yield of lentil genotypes in fall planting were more than 4 and about 2 times of spring planting, respectively. In the 2008/09 growing seasons 39% of spring planting samples produced less than 50 gr/m² seed yield. It seems that the improvement of seed yield in the fall planting were due to suitable winter survival of plants fallowed by better usage of rainfall and escaping from drought and heat stress of the end of spring. Results showed that there were some cold tolerant accessions in MLC and it is possible to provide cold tolerant cultivars for fall planting in Mashhad conditions.     

Sperm of feral carp <Emphasis Type="Italic">Cyprinus carpio</Emphasis>: optimization of activation solution

The spermatozoa of oviparous fish, such as feral carp (Cyprinus carpio), are immotile in the presence of semen plasma or isotonic solutions, and to obtain good motility, they must be diluted with suitable medium. The objective of this study was to identify the best activating solution for feral carp sperm. Sperm motilities were compared in the new activating solution (a): (50 mM NaCl, 30 mM KCl, 30 mM Tris, pH = 8.5) and activating solution (b): (50 mM NaCl, 40 mM KCl, 30 mM Tris, pH = 8.5) based on effect of pH with everyone of Na⁺ and K⁺ ions versus four other activating solutions Billard’s saline solution, Poupard’s saline solution, distilled water and hatchery water that is routinely used for extending carp semen. Our results showed that maximum total motility period and percentage of motile sperm were seen in selected saline solution (a). The present study describes an activating solution that prolongs feral carp sperm motility.     

Inotropic and chronotropic effects of 6-hydroxy-4-methylquinolin-2(1H)-one derivatives in isolated rat atria

Background: Selective phosphodiesterase (PDE3) inhibitors improve cardiac contractility and may use in congestive heart failure. However, their proarrhythmic potential is the most important side effect. Methods: In this research, we evaluated the potential cardiotonic activity of six new synthesized selective PDE3 inhibitors (6-hydroxy-4-methylquinolin-2(1H)-one derivatives) using the spontaneously beating atria model. In each experiment, atrium of reserpine-treated rat was isolated and the contractile and chronotropic effects of a synthesized compound were assessed. The 3-isobutyl-1-methylxanthine, a non-selective PDE inhibitor, was used for comparison. Results: The results showed that, among new compounds, the best pharmacological profile was obtained with the compound 6-[4-(4-methylpiperazine-1-yl)-4-oxobutoxy]-4-methylquinolin-2(1H)-one, C6, which displayed selectivity for increasing the force of contraction (168 +/- 5% change over the control) rather than the frequency rate (138 +/- 5% change over the control) at 300 muM. However, C6 at concentrations of 10 and 100 muM produced left and upward shift in the positive inotropic concentration-response curve of isoprenaline. The -log EC50 of isoprenaline was 8.843 +/- 0.171 in the absence, 9.448 +/- 0.138 and 9.456 +/- 0.107 in the presence of 10, 100 muM of C6, respectively (P<0.001, n = 6). Also, amrinone, a selective PDE3 inhibitor, shifted the isoprenaline concentration-response curve to the left and upward. The concentration of 10 and 100 muM amrinone decreased -log EC50 of isoprenaline to 9.527 +/- 0.287 and 9.423 +/- 0.243, respectively (P<0.001, n = 6). Moreover, the positive chronotropic effect of isoprenaline was not affected by amrinone or C6. Conclusion: This study provides functional evidence for the positive inotropic effect of C6. Considering the augmentation of isoprenaline positive inotropic concentration-response with C6 and amrinone, we conclude that C6 produces its effect via potentiation of cAMP-dependent signaling system and possibly by inhibition of PDE3 activity.     

Prevalence of avian influenza (H9N2) in commercial quail,partridge, and turkey farms in Iran, 2014–2015

Tropical Animal Health and Production - Avian influenza virus (AIV) H9N2 subtype is endemic in Iran and causes substantial economic loss to the growing poultry industry within the country. In this...     

Vaccination in Leishmaniasis: A Review Article

Leishmaniasis is caused by protozoan Leishmania parasites that are transmitted through female sandfly bites. The disease is predominantly endemic to the tropics and semi-tropics and has been reported in more than 98 countries. Due to the side effects of anti-Leishmania drugs and the emergence of drug-resistant isolates, there is currently no encouraging prospect of introducing an effective therapy for the disease. Hence, it seems that the key to disease control management is the introduction of an effective vaccine, particularly against its cutaneous form. Advances in understanding underlying immune mechanisms are feasibale using a variety of candidate antigens, including attenuated live parasites, crude antigens, pure or recombinant Leishmania proteins, Leishmania genes encoding protective proteins, as well as immune system activators from the saliva of parasite vectors. However, there is still no vaccine against different types of human leishmaniasis. In this study, we review the works conducted or being performed in this field. Key Words: Immune response, Leishmaniasis, Vaccination     

Association of tcdA+/tcdB+ Clostridium difficile Genotype with Emergence of Multidrug-Resistant Strains Conferring Metronidazole Resistant Phenotype

Background:

Reduced susceptibility of Clostridium difficile to antibiotics is problematic in clinical settings. There is new evidence indicating the cotransfer of toxin-encoding genes and conjugative transposons encoding resistance to antibiotics among different C. difficile strains. To analyze this association, in the current study, we evaluated the frequency of toxigenic C. difficile among the strains with different multidrug-resistant (MDR) profiles in Iran.

Methods:

Antimicrobial susceptibility patterns and minimal inhibitory concentrations (MIC) of the isolates were determined against metronidazole, imipenem, ceftazidime, amikacin, and ciprofloxacin by agar dilution method. The association of the resistance profiles and toxigenicity of the strains were studied by PCR targeting tcdA and tcdB genes.

Results:

Among 86 characterized strains, the highest and lowest resistance rates were related to ciprofloxacin (97%) and metronidazole (5%), respectively. The frequency of resistance to other antibiotics was as follow: imipenem (48%), ceftazidime (76%), and amikacin (76.5%). Among the resistant strains, double drug resistance and MDR phenotypes were detected in the frequencies of 10.4% and 66.2%, respectively. All of the metronidazole-resistant strains belonged to tcdA ⁺/tcdB ⁺ genotype with triple or quintuple drug resistance phenotypes. MIC₅₀ and MIC₉₀ for this antibiotic was equally ≤ 8 μg/ml.

Conclusion:

These results proposed the association of tcdA ⁺/tcdB ⁺ genotype of C. difficile and the emergence of resistance strains to broad-spectrum antibiotics and metronidazole. Key Words: Multidrug resistance, Clostridium difficile, Metronidazole     

Neuronal Cell Reconstruction with Umbilical Cord Blood Cells in the Brain Hypoxia-Ischemia

Background: Brain hypoxia-ischemia is a human neonatal injury that is considered a candidate for stem cell therapy. Methods: The possible therapeutic potential of human umbilical cord blood (HUCB) stem cells was evaluated in 14-day-old rats subjected to the right common carotid occlusion, a model of neonatal brain hypoxia-ischemia. Seven days after hypoxia-ischemia, rats received either saline solution or 4 × 10⁵ HUCB cells i.v. Rats in control group did not receive any injection. After two weeks, rats were assessed using two motor tests. Subsequently, rats were scarified for histological and immunohistochemical analyses. Results: Our immunohistochemical findings demonstrated selective migration of the injected HUCB cells to the ischemic area as well as reduction in infarct volume. Seven days after surgery, we found significant recovery in the behavioral performance in the test group (12.7 +/- 0.3) compared to the sham group (10.0 +/-0.05), a trend which continued to day 14 (15.3 ± 0.3 vs. 11.9 ± 0.5, P<0.05). Postural and motor asymmetries at days 7 and 14 in the test group showed a significant decrease in the percentage of right turns in comparison to the sham group (75% and 59% vs. 97% and 96%, P<0.05). Conclusion: The results show the potential of HUCB stem cells in reduction of neurologic deficits associated with neonatal hypoxia-ischemia. Key Words: Hypoxia-Ischemia, Nerve cell, Umbilical Cord Blood     

Inhibitory effects of Sage extract on the growth of enteric bacteria

Antibacterial activity of Sage extract at concentrations of 0.1, 0.05, 0.025, 0.0125, 0.00625, 0.003125, 0.00156, 0.0005 and 0.00025 g dL(-1) against Salmonella typhi, Shigella sonnei, S. flexneri, Proteus vulgaris, Staphylococcus aureus, ETEC Escherichia coli and Pseudomonas aeruginosa was evaluated. Susceptibility testing of bacterial strains against 18 antibiotics was also performed for comparison. The results showed that P. aeruginosa and ETEC E. coli were completely resistant to Sage extract even at concentration of 0.1 g dL(-1). Its antibacterial activity (0.1 g dL(-1)) against P. vulgaris, S. flexneri and S. sonnei was the same as nitrofurantoin and ampicilline respectively. Sage extract (0.1 and 0.05 g dL(-1)) exhibited the same effects as ampicilline and streptomycin against S. typhi. Its antibacterial activity (0.1, 0.05 and 0.25 g dL(-1)) against S. aureus was the same as ceftazidim, chloramphenicol, gentamycin, neomycin and nitrofurantoin and was more significant compared to streptomycin and vancomycin. The results suggest Sage can be considered as an alternative herbal in the treatment of infections caused by the above-mentioned bacteria.     

Beta cell protective effects of sodium tungstate in streptozotocin-induced diabetic rats: glycemic control, blockage of oxidative stress and beta cell histochemistry

Background: Diabetes is a major public health problem. The development of new therapies that are able to improve glycemia management and even to cure diabetes is of great interest. In this study, protective effects of sodium tungstate against STZ-induced beta-cell damages were investigated. Methods: Sixty rats were divided into six groups: control, diabetic, sodium tungstate treated diabetic rats from one week before STZ injection (TDB), food-restricted diabetic (FRD), tungstate treated control, sodium tungstate treated diabetic rats from one week after STZ administration (TDA). We evaluated serum insulin, glucose and glucose tolerance; liver glycogen content, glucokinase (GK) activity; blood and pancreas antioxidant power, lipid peroxidation; and fuchsin-aldehyde histochemical staining of beta-cells. Results: Blood glucose levels of TDB group were lower than other diabetic groups (P<0.01). Blood insulin levels of all diabetic groups were lower than controls (P<0.01). Glucose intolerance improved in TDB animals. Blood and pancreas antioxidant power, liver glycogen contents and GK activities and granulated beta cells increased in TDB rats in comparison with other diabetic groups (P<0.01). Likewise, lipid peroxidation decreased significantly in TDB rats (P<0.01). Conclusions: Results suggested that sodium tungstate if administrated before STZ injection improves glycemic state by a direct effect on pancreatic beta-cells and preserves them by reducing the activity of these cells at the time of STZ injection, reducing STZ-induced oxidative stress, reducing insulin secretion, or all of the above mentioned.