In vitro effects of bacterial lipopolysaccharide (LPS) on motion parameters of ram epididymal sperm

This study investigated in vitro effects of bacterial lipopolysaccharide (LPS) on ram sperm motion parameters at concentrations less than its spermicidal effects. After incubation of epididymal sperm samples for 45, 90, 135, 180, and 225 min in the presence of LPS (50, 100, 200, and 400 μg/ml), motion parameters were evaluated by computer-aided sperm analysis. There was no significant increase of the fast progressive motility (class A) of sperm in LPS-treated groups at different times while there was a decrease in the slow progressive motility (class B) and the non-progressive motility (class C) of sperm in LPS-treated groups at different times that were only significant (P < 0.05) at the 135 min (classes B and C) and 180 min (only class C) incubation times and at concentrations of 200 and 400 μg/ml LPS compared to their controls. Concentrations of 100, 200, and 400 μg/ml LPS caused an increase in linearity; the wobble, mean angular displacement, and the amplitude of lateral head displacement decreased significantly (P < 0.05) at different times (45, 90, 135, 180, and 225 min). It can be concluded that LPS, at concentrations less than its spermicidal effect, changes progressive motility and motion parameters that probably lead to infertility.     

The effect of verapamil on in vitro susceptibility of promastigote and amastigote stages of <Emphasis Type="Italic">Leishmania tropica</Emphasis> to meglumine antimoniate

Pentavalent antimonials are the standard treatment for cutaneous leishmaniasis (CL) with low efficacy and resistance is emerging. CL is increased significantly in respect to incidence rate and expanding to new foci. In the present study, the effect of verapamil on in vitro susceptibility of promastigote and amastigote stages of Leishmania tropica to meglumine antimoniate (MA, Glucantime) was evaluated using colorimetric assay (MTT) and in a macrophage model, respectively. Verapamil, as a calcium channel blocker, affects drug uptake by preventing of drug efflux from the cells. In promastigote form, several concentrations of MA with or without verapamil showed significant decrease (P < 0.05) in optical density. The overall mean IC₅₀ value with combination of MA plus verapamil (IC50 = 116.03 μg/ml) was significantly less than MA (IC50 = 225.14 μg/ml) alone (P < 0.05) for promastigote stage. Similarly, the amastigote stage was more susceptible to treatment with MA plus verapamil to that of MA alone (P < 0.05). Analysis of overall effect of different concentrations of MA alone, compared with combination of MA plus verapamil by mean infection rate of amastigotes in each macrophage showed a significant difference (P < 0.05).These findings indicated some degree of synergistic effects between MA and verapamil on in vitro susceptibility of L. tropica to MA. Further works are required to evaluate this synergistic effect on animal model or volunteer human subjects.     

In vitro and in vivo antileishmanial efficacy of nitazoxanide against Leishmania donovani

Control of Leishmania infection relies primarily on chemotherapy, and the current drug available for treating leishmaniasis is limited. Nitazoxanide (NTZ) is a broad spectrum antiparasitic agent with activity against protozoa, nematodes, cestodes, and trematodes. In the present study, the in vitro antileishmanial efficacy of NTZ was evaluated by incubation of Leishmania donovani promastigotes with NTZ, indicating that NTZ can affect the ultrastructure of parasite promastigote and efficiently inhibit the parasite growth. Moreover, 200 μg/ml NTZ inhibited >90% of promastigotes growth, showing similar activity of the reference drug amphotericin B (P > 0.05). Therapeutic efficacy of NTZ against L. donovani-infected BALB/c mice demonstrated that oral NTZ produced a significant reduction of parasite burden in spleen and liver from L. donovani-infected mice, compared with the untreated mice (P < 0.05). These results indicated NTZ may be a novel therapeutic drug for leishmaniasis.     

Comparative studies of the anti-leishmanial activity of three <Emphasis Type="Italic">Crotalus durissus</Emphasis> ssp. venoms

In this study, we compared the anti-leishmanial activity of three crotalic venoms (Crotalus durissus terrificus–Cdt, Crotalus durissus cascavella–Cdca, and Crotalus durissus collilineatus–Cdcol). Different concentrations of each venom incubated with Leishmania (Leishmania) amazonensis promastigotes were used. Cdt venom exhibited a higher anti-leishmanial activity (Inhibitory concentration-IC50-value of 4.70 ± 1.72 μg/ml) in comparison with that of Cdca venom (IC50 value of 9.41 ± 1.21 μg/ml), while Cdcol venom increased parasite numbers in 50% at a concentration of 44.30 ± 2.18 μg/ml. In addition, this venom showed a low anti-leishmanial activity in higher concentrations (IC50 value of 281.00 ± 9.50 μg/ml). The main fractions of Cdca venom were isolated and assayed under similar conditions used for assessing crude venom. The most active fractions were gyroxin and crotamine that had IC50 values of 3.80 ± 0.52 μg/ml and 19.95 ± 4.21 μg/ml, respectively. Convulxin also inhibited parasite growth rate, although this effect was not dose-dependent. Crotoxin was the least effective fraction with an IC50 value of 99.80 ± 2.21 μg/ml. None of the protein fractions presented cytotoxic effects against J774 cells in culture. In vivo assays using BALB/c mice revealed that crotoxin and crotamine were the main toxic fractions. In conclusion, C. durissus cascavella venom has three main fractions with anti-leishmanial activity. These results open new possibilities to find proteins that might be used as possible agents against cutaneous leishmaniasis.     

In vitro anthelmintic activity of the essential oils of <Emphasis Type="BoldItalic">Zanthoxylum zanthoxyloides</Emphasis> and <Emphasis Type="BoldItalic">Newbouldia laevis</Emphasis> against <Emphasis Type="BoldItalic">Strongyloides ratti</Emphasis>

The need for new anthelmintic with no chemical residues is becoming urgent. In a program aiming at the evaluation of plant as sources of new active molecules, the anthelmintic activities of the essential oils (EOs) obtained from either Zanthoxylum zanthoxyloides seeds or Newbouldia laevis leaves were evaluated against Strongyloides ratti by analyzing the results of two in vitro bioassays. These two plants and their tested parts were retained after an ethnopharmacology survey that confirmed their use by small-scale farmers for treatment of small ruminants affected by digestive helminths. The plants were harvested in Benin, and their EO were obtained by hydrodistillation. The EO yield of extraction was 0.65% (w/w) of for Z. zanthoxyloides seeds and 0.05% (w/w) for N. laevis. The chemical compositions of the two EOs were analyzed by gas chromatography coupled with mass spectrometry. The major constituents of the EO from Z. zanthoxyloides consisted of the following compounds: γ-terpinene (18 %), undecane (15 %), valencene (8.3 %), decanal (8.3 %), and 3-carene (6.7 %). In contrast, the major constituents of the EO from N. laevis leaves consisted of the following compounds: β-caryophyllene (36 %) and eugenol (5.8 %). An egg-hatching inhibition (EHI) assay was developed and a larval migration inhibition assay was used on S. ratti to examine the effects of the EOs and to evidence their inhibitory concentrations (IC₅₀ and IC₉₀) values on this nematode. Furthermore, the toxicity of the two EOs on Vero cell line was evaluated. When tested on S. ratti egg hatching, the two EOs resulted in similar IC₅₀ values (19.5 and 18.2 μg/ml for Z. zanthoxyloides and N. laevis, respectively), which were about sevenfold higher than that of the control (thiabendazole, IC₅₀ = 2.5 μg/ml). Larval migration was inhibited at similar concentrations for: Z. zanthoxyloides (IC₅₀ = 46 μg/ml), N. laevis (IC₅₀ = 51 μg/ml), and the control [levamisole (IC₅₀ = 36 μg/ml)]. No cytotoxicity was found on Vero cells because both EOs had IC₅₀ values higher than 50 μg/ml. Therefore, we have concluded that the EOs from two plants, used in folk medicine, may contain compounds with anthelmintic activity and could be used as improved traditional medicines or, at least, as food additives in a combined treatment for the control of helminth infections.     

Screening of antiplasmodial efficacy of <Emphasis Type="Italic">Ajuga bracteosa</Emphasis> Wall ex. Benth

The rising problem of Plasmodium resistance to the classical antimalarial drugs stresses the need to look for newer antiplasmodial components with effective and new mode of action. In the present study, the traditional medicinal plant Ajuga bracteosa has been screened for its antiplasmodial efficacy. The extract was found to possess significant in vitro antiplasmodial efficacy with an IC₅₀ of 10.0 μg/ml. Thus, the extract was further evaluated for its in vivo schizontocidal activity and efficacy in terms of survival time in Plasmodium berghei infected BALB/c mice. The extract at 250, 500, and 750 mg/kg/day exhibited significant (p < 0.0001) blood schizontocidal activity during established infection with enhanced mean survival time comparable to that of standard drug chloroquine, 5 mg/kg/day. The significant schizontocidal activity and enhanced mean survival time of mice stress the need to identify and characterize active antiplasmodial principle from this plant.     

Clinical features and treatment outcomes of vancomycin-intermediate <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> (VISA) and heteroresistant vancomycin-intermediate <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> (hVISA) in a tertiary care institution in Singapore

This retrospective case–control study was undertaken to review the clinical features associated with heteroresistant vancomycin-intermediate Staphylococcus aureus (hVISA) and vancomycin-intermediate S. aureus (VISA) infections and the local impact they have on clinical outcome. Compared with vancomycin-susceptible S. aureus (n = 30), hVISA and VISA infections (n = 10) are found to be associated with a longer period of prior glycopeptide use (P = 0.01), bone/joint (P < 0.01) and prosthetic infections (P = 0.04), as well as treatment failure, as evidenced by longer bacteremic (P < 0.01) and culture positivity (P < 0.01) periods. This was observed to have resulted in longer hospital length of stay (P < 0.01) and total antibiotic therapy duration (P = 0.01). There was, however, no significant difference in the overall patient mortality or the hospitalization cost (P = 0.12) in both groups. Clinicians should be cognizant of the association between hVISA/VISA with high bacterial load deep-seated infections. We recommend targeted and even universal screening for hVISA/VISA in methicillin-resistant S. aureus (MRSA) infections.     

RNAi-mediated silencing of a novel Ascaris suum gene expression in infective larvae

In the present study, the potential of RNA interference (RNAi) as a gene silencing tool and the resultant effects on Ascaris suum larval development was examined by targeting a gene (represented by the EST 06G09) specifically expressed in the infective larvae of A. suum. BALB/c mice were infected with RNAi-treated larvae. The results showed that the target gene was silenced after soaking for 72 h, and the survival rate of the RNAi-treated larvae was reduced by 17.25% (P < 0.01). A significant difference (P < 0.05) was detected in the numbers of larvae collected from the livers and lungs of infected mice 4 days after infection with untreated larvae (164.29 ± 21.51) and RNAi-treated larvae (71.43 ± 14.35). Significant differences (P < 0.01) were also found in the body length and width between untreated larvae (480 ± 105.77 μm for length and 23.93 ± 3.72 μm for width) and RNAi-treated larvae (400.57 ± 71.31 μm for length and 20.20 ± 2.43 μm for width). These results show that the gene represented by EST 06G09 may play a role in the development of A. suum larvae.     

Higher Doses of Subcutaneous IgG Reduce Resource Utilization in Patients with Primary Immunodeficiency

The recommended dose of IgG in primary immunodeficiency (PID) has been increasing since its first use. This study aimed to determine if higher subcutaneous IgG doses resulted in improved patient outcomes by comparing results from two parallel clinical studies with similar design. One patient cohort received subcutaneous IgG doses that were 1.5 times higher than their previous intravenous doses (mean 213 mg/kg/week), whereas the other cohort received doses identical to previous subcutaneous or intravenous doses (mean 120 mg/kg/week). While neither cohort had any serious infections, the cohort maintained on higher mean IgG dose had significantly lower rates of non-serious infections (2.76 vs. 5.18 episodes/year, P < 0.0001), hospitalization (0.20 vs. 3.48 days/year, P < 0.0001), antibiotic use (48.50 vs. 72.75 days/year, P < 0.001), and missed work/school activity (2.10 vs. 8.00 days/year, P < 0.001). The higher-dose cohort had lower health care utilization and improved indices of well being compared to the cohort treated with traditional IgG doses.     

New Colorimetric Microdilution Method for In Vitro Susceptibility Testing of Borrelia burgdorferi Against Antimicrobial Substances

 A newly developed colorimetric microdilution method was used to analyze the activity of 12 antimicrobial agents against nine Borrelia burgdorferi isolates, including all three genospecies pathogenic for humans. In addition, in vitro antimicrobial resistance patterns of Borrelia valaisiana and Borrelia bissettii tick isolates were investigated. The applied test system is based upon color changes that occur in the presence of phenol red and result from the accumulation of nonvolatile acid produced by actively metabolizing spirochetes. After 72 h of incubation, minimal inhibitory concentrations (MICs) were determined from the decrease of absorbance by software-assisted calculation of growth curves. MIC values were lowest for azlocillin (MIC, ≤0.125 μg/ml), ceftriaxone (MIC range, ≤0.015–0.06 μg/ml), and azithromycin (MIC range, ≤0.015–0.06 μg/ml). Whereas tobramycin (MIC range, 8–64 μg/ml) exhibited little activity, spectinomycin (MIC range, 0.25–2 μg/ml) showed in vitro antimicrobial activity against Borrelia burgdorferi. The MICs of penicillin G for Borrelia afzelii isolates were ten times higher than those for Borrelia burgdorferi, Borrelia valaisiana, and Borrelia bissettii isolates (P<0.05) and 100 times higher than those for isolates belonging to the genospecies Borrelia garinii (P<0.05). Further significant differences with respect to the MIC values of the other antimicrobial agents tested were not noted. The colorimetric microdilution method offered the advantages of reliability, reproducibility, and convenience and could handle large numbers of isolates and antibiotics.     

Determinants for the Development of Oropharyngeal Colonization or Infection by Fluconazole-Resistant Candida Strains in HIV-Infected Patients

 A point prevalence study to document oral yeast carriage was undertaken. Risk factors for the development of oropharyngeal colonization or infection by fluconazole-resistant Candida strains in HIV-infected patients were investigated with a case-control design. Cases included all patients with fluconazole-resistant strains (MIC≥64 μg/ml), and controls were those with susceptible (MIC≤8 μg/ml) or susceptible-dependent-upon-dose (MIC 16–32 μg/ml) strains. One hundred sixty-eight Candida strains were isolated from 153 (88%) patients, 28 (16%) of whom had oropharyngeal candidiasis. Overall, 19 (12%) of the patients harbored at least one resistant organism (MIC≥64 μg/ml). Among patients with resistant strains, tuberculosis (P<0.001), esophageal candidiasis (P=0.001), clinical thrush (P<0.001), and a CD4+ cell count <200/mm³ (P=0.03) were more frequent. These patients had also been treated more commonly with antituberculous drugs (adjusted odds ratio [OR] 6.13; 95% confidence interval [CI] 2.11–17.80), ciprofloxacin (OR 6.0; 95% CI 1.23–29.26), fluconazole (OR 4.59; 95% CI 1.55–13.52), and steroids (OR 4.13; 95% CI 1.11–15.39). Multivariate analysis showed that the determinants for fluconazole resistance were therapy with antituberculous drugs (OR 3.61; 95% CI 1.08–12.07;P=0.03) and one of the following: previous tuberculosis (OR 3.53; 95% CI 1.08–14.57;P=0.03) or fluconazole exposure (OR 3.41; 95% CI 1.10–10.54). Findings from this study indicate that treatment with antituberculous drugs, previous tuberculosis, and fluconazole exposure are the strongest determinants for development of oropharyngeal colonization or infection by fluconazole-resistant Candida strains in HIV-infected patients.     

Effect of xylazine–ketamine–diazepam anesthesia on certain clinical and arterial blood gas parameters in sheep and goats

This study was conducted to evaluate the effect of xylazine–ketamine–diazepam anesthesia on heart rate, respiration rate, rectal temperature, rumen motility, peripheral blood pH, PaO2, and PaCO2 in adult female nonpregnant Awassi sheep and adult female nonpregnant Damascus goats. Anesthesia was induced using 0.1 mg/kg, 5 mg/kg, and 0.25 mg/kg xylazine, ketamine, and diazepam respectively as a single intravenous injection. The heart rate, respiration rate, rectal temperature, rumen motility, peripheral arterial blood pH, PaO2, and PaCO2 were evaluated 15 min before and at 15, 30, and 60 min during anesthesia. In sheep, the heart rate, rumen motility, and PaO2 were decreased significantly (P < 0.05) at 15, 30, and 60 min following anesthesia. The respiration rate and rectal temperature and blood pH were decreased significantly (P < 0.05) at 30 and 60 min. The peripheral PaCO2 was increased significantly (P < 0.05) at 15 and 30 min. In goats, the heart rate and rumen motility were decreased significantly (P < 0.05) at 15, 30, and 60 min while the respiration rate was decreased only significantly (P < 0.05) at 60 min. Rectal temperature was decreased significantly (P < 0.05) at 30 and 60 min. The blood pH was decreased significantly (P < 0.05) at 15 and 30 min. PaO2 was only significantly (P < 0.05) decreased at 15 min while PaCO2 was increased significantly (P < 0.05) at 15 and 30 min.     

Efficacy of tigecycline vs. imipenem in the treatment of experimental Acinetobacter baumannii murine pneumonia

The in vivo activity of tigecycline was evaluated in an experimental pneumonia model (C57BL/6 mice) by Acinetobacter baumannii. Two clinical strains were used: minimum inhibitory concentrations (MICs) of imipenem and tigecycline 1 and 2 μg/mL (imipenem-susceptible, IPM-S), and 8 and 2 μg/mL (imipenem-intermediate, IPM-I), respectively. For imipenem (30 mg/Kg), ∆T/CMI (h) were 1.04 and 0.51 for IPM-S and IPM-I, respectively. For tigecycline (5 mg/Kg), the area under the concentration–time curve (AUC)/MIC_0–24 h (serum and lung) were 9.24 and 4.37 (for the two strains), respectively. In the efficacy experiments with the IPM-S, imipenem (log CFU/g 3.59 ± 0.78, p = 0.006) and tigecycline (2.82 ± 1.2, p = 0.054) decreased the bacterial counts in lungs with respect to its controls; with the IPM-I, both imipenem (1.21 ± 0.52, p = 0.002) and tigecycline (3.21 ± 0.28, p = 0.035) decreased the bacterial counts with respect to the controls. In the survival experiments, with the IPM-S, the mortality was the same in the control (67%) and in the tigecycline (77%) groups, and imipenem reduced it (21%, p = 0.025); with the IPM-I, the mortality was the same in the control (87%) and in the tigecycline (85%) groups, and imipenem (0%) reduced it (p < 0.001). In summary, the present study shows that tigecycline is less efficacious than imipenem in the treatment of experimental A. baumannii pneumonia caused by IPM-S and IPM-I strains.     

Influence of sympatho-vagal balance on airway responsiveness in athletes

The increased prevalence of airway hyper-responsiveness (AHR) observed among athletes suggests that high-level training may contribute to the development of AHR. We investigated the possible influence of the sympatho-vagal balance on this phenomenon in 40 athletes and 10 sedentary controls. Each subject filled out a respiratory questionnaire, had a methacholine challenge, and measurements were made of their baseline plasma catecholamines [epinephrine (E), norepinephrine (NE) and dopamine (DA)] as a reflection of sympathetic tone, and their heart rate variability (SDNN: standard deviation of all normal-to-normal intervals) as an indicator of parasympathetic tone. The athletes had a 45% prevalence of AHR (defined as PC₂₀ < 16 mg/ml, where PC₂₀ is the concentration of methacholine inducing a 20% fall in the forced expiratory volume in 1 s, FEV₁) with a mean PC₂₀ of 21.2 mg/ml compared with 10% prevalence (mean PC₂₀: 74.4 mg/ml) in sedentary subjects (P < 0.01). Plasma catecholamine values were not significantly different between the two groups (all P > 0.05), but the estimated parasympathetic tone was higher in athletes (P=0.01). When data from all subjects were analyzed together, plasma E and NE correlated with PC₂₀ (r=0.39, P=0.005 and r=0.29, P < 0.005) but DA and SDNN did not (both P > 0.05). However, the ratios E/SDNN, NE/SDNN and DA/SDNN showed significant correlations with PC₂₀ (r=0.42, P < 0.01; r=0.33, P < 0.005 and r=0.31, P < 0.05, respectively). This study suggests that the sympatho-vagal balance may contribute to the increased AHR in the population studied but this influence alone cannot explain the higher prevalence of AHR in athletes. Accepted: 26 July 2000     

Antibody decay after immunisation of health-care workers with an acellular pertussis vaccine

Antibody decay after a single dose of acellular pertussis vaccine containing 25 μg of pertussis toxin (PT), 25 μg of filamentous hemagglutin (FHA), and traces of pertactin (PRN) was monitored in health-care workers. Blood was sampled 4 weeks (n = 246), 1 year (n = 187), 2 years (n = 53), 3 years (n = 134), and 4 years (n = 37) after vaccination. IgG anti-PT, IgG anti-FHA and IgG anti-PRN were measured by ELISA. Peak median antibodies to PT, FHA, and PRN were 314, 785, and 84 EU/ml respectively. IgG anti-PT decreased to a median of 29% (76 EU/ml), 18% (64 EU/ml), 19% (58 EU/ml), and 20% (63 EU/ml) of the peak value after 1, 2, 3, and 4 years respectively. IgG anti-FHA decreased more slowly, but showed similar decay patterns. In German health-care workers antibodies to pertussis antigens decayed rapidly within the first year after vaccination, but remained stable after 2, 3, and 4 years. This observation may have implications for the timing of booster vaccinations in adults. Parts of this study were presented as a poster (Poster #129) during the 8th International Symposium “Saga of the genus Bordetella,” 7–10 November 2006 at the Institut Pasteur in Paris, France.     

In vivo validation of Aloe ferox (Mill). Elephantorrhiza elephantina Bruch. Skeels. and Leonotis leonurus (L) R. BR as potential anthelminthics and antiprotozoals against mixed infections of gastrointestinal nematodes in goats

Aloe ferox (Mill)., Elephantorrhiza elephantina Bruch. Skeels. and Leonotis leonurus (L) R. BR. are some of the plants used by farmers in the Eastern Cape Province to control worms in goats, but information on their efficacy is lacking. The study was conducted to determine efficacy of these plants on gastrointestinal nematodes in natural mixed infections in goats. Forty-eight male goats aged 8–12 months were divided into eight groups (Treatments A–H) of six animals each, balanced in terms of liveweight and worm egg count. Treatments A to F received plant extracts, three animals in each group receiving doses of 250 mg/kg and the other three receiving 500 mg/kg at concentration of 100 mg/ml, while those in G and H received Valbazen? (11.36% albendazole) at 10 mg/kg, and 0.5 ml/kg distilled water, respectively per os. Faecal samples were collected on days 0, 3, 6 and 9 for faecal egg counts (FEC), and body weights recorded on days 1 and 9. Results showed significant reductions (P < 0.05) in strongyle eggs by A. ferox extract at dose levels of 500 mg/kg on days 3, 6 and 9, while reductions in Eimeria spp. oocysts were observed on days 3, 6 and 9 for animals that received 500 mg/kg doses. E. elephantina caused significant reduction (P < 0.05) of Trichuris spp. eggs on days 3 and 6, respectively at 250 mg/kg dose level, whereas L. leonurus also caused significant reduction (P < 0.05) in FEC of Trichuris spp. and Eimeria spp. oocysts at 250 mg/kg dose level on day 9. Albendazole caused reductions (P < 0.05) in strongyle eggs on days 3 and 6, Trichuris spp. on days 3, 6 and 9, and on coccidia, it caused a reduction (P > 0.05) on day 1, whereas on days 6 and 9, there was an increase. On total mixed infections, highest FECR% were observed with the extract of A. ferox on days 3 (53%), 6 (54%) and 9 (58%) at 500 mg/kg,whereas albendazole had efficacy levels of 39%, 44% and 29% on days 3, 6 and 9, respectively. Body weight of goats from days 1 to 9 were not significant different from the control. The study revealed efficacy of A. ferox, E. elephantina and L. leonurus against gastrointestinal parasites at high doses (500 mg/kg), showing that the plants have the potential to be used as anthelminthics.     

Relationship between plasma thyroid hormones and some biochemical parameters in Iranian Sarabi calves

The thyroid gland has some important endocrine hormones that regulate basal metabolism in various tissues of domestic animals. Thyroid hormones have a central role in animals’ development and their tissue functions. In this study, the relationship between the plasma thyroxine (T₄), triiodothyronine (T₃), free thyroxine (fT₄), free triiodothyronine (fT₃), triglyceride, cholesterol, glucose, total protein, and albumin concentrations as well as albumin/globulin ratio in different ages of Iranian Sarabi calves was investigated. Blood samples were collected from the jugular vein of 47 clinically healthy calves free from internal and external parasites (grouped according to their age—1–14 days, 1–2, and 3–6 months) in early of winter. The level of thyroid hormones was determined by chemiluminescence, and other parameters were measured by spectrophotometry using commercial kits. Our data from this study indicates that there was no significant difference and correlation in all the studied parameters between age groups and sexes. But we found a significant correlation between plasma T₄ and total protein (P < 0.05, r = 0.600), T₄ and albumin (P < 0.05, r = 0.575), T₃ and fT₃ (P < 0.05, r = 0.610), T₃ and total protein (P < 0.01, r = 0.725), T₃ and glucose (P < 0.01, r = 0.685), and fT₄ and fT₃ (P < 0.05, r = 0.609) concentrations as well as between total protein and albumin/globulin ratio (P < 0.01, r = −0.783).     

Cloning,expression, and characterization of <Emphasis Type="Italic">Schistosoma japonicum</Emphasis> tegument protein phosphodiesterase-5

The tegument proteins of schistosomes are regarded as potential vaccine candidates and drug targets to control schistosomiasis. Nucleotide pyrophosphatase/phosphodiesterase-5 (NPP-5), which belongs to a multigene family of nucleotide pyrophosphatase/phosphodiesterases (NPPs), is important in the hydrolysis of pyrophosphate or phosphodiester bonds in nucleotides and their derivatives. In the present study, SjNPP-5, identified as one of the tegument proteins of Schistosoma japonicum in our previous proteomic studies, was cloned on a fragment of 1,371 bp and expressed as a recombinant protein of 69 kDa. Real-time RT-PCR analysis showed that SjNPP-5 was up-regulated at 21–42 days, and the expression level in 42-day-old male worms was almost nine times higher than that in females. Western blot analysis revealed that rSjNPP-5 had good antigenicity. Immunofluorescence analysis found that SjNPP-5 was a membrane-associated antigen mainly distributed on the surface of the male adult worm of S. japonicum. BALB/c mice vaccinated with rSjNPP-5 three times showed a 29.90% worm reduction (P < 0.05) and a 26.21% egg count reduction (P > 0.05). Immunization with rSjNPP-5 induced a mixed Th1/Th2 response in which Th1 was dominant. The response was characterized by a reduced IgG1/IgG2a ratio and elevated production of cytokines IFN-γ and IL-4. This study suggested that SjNPP-5 may be important in schistosome development, and further investigations are required to fully understand the function of this molecule.     

4-epi-Pimaric acid: a phytomolecule as a potent antibacterial and anti-biofilm agent for oral cavity pathogens

The present study focused on the antibacterial and biofilm inhibitory potential of 4-epi-pimaric acid isolated from aerial parts (stem and leaves) of Aralia cachemirica L. (Araliaceae) against oral cavity pathogens. 4-epi-Pimaric acid exhibited minimum inhibitory concentration (MIC) in the range of 4–16 μg/ml and minimum bactericidal concentration (MBC) two- to four-folds higher than MIC. There was significant inhibition in the biofilm formation by Streptococcus mutans on the saliva coated surface (P < 0.05), and confocal microscopy revealed that 4-epi-pimaric acid inhibited the clumping and attachment of S. mutans. At 8 × MIC concentration, it significantly prevented the pH drop and reduced S. mutans biofilms (P < 0.05). Increased propidium iodide staining and leakage of 260- and 280-nm absorbing material by 4-epi-pimaric acid treated cells of S. mutans suggested that it probably causes disruption of the cytoplasmic membrane structure. It also exhibited significant suppression of TNF-α expression in human neutrophils, suggestive of its anti-inflammatory activity. Furthermore, the compound was found to be significantly safe (IC₅₀ >100 μg/ml) in the MTT assay on AML-12 cell lines. In conclusion, 4-epi-pimaric acid showed promising antibacterial, anti-biofilm and anti-inflammatory potency and this compound can be exploited for therapeutic application in oral microbial infections.     

In vitro antiplasmodial activity and cytotoxicity of crude extracts and compounds from the stem bark of <Emphasis Type="Italic">Kigelia africana</Emphasis> (Lam.) Benth (Bignoniaceae)

In order to assess the potential of the stem bark of Kigelia africana (Lam.) Benth as source of new anti-malarial leads, n-hexane and ethyl acetate (EtOAc) extracts and four compounds isolated from the stem bark were screened in vitro against the chloroquine-resistant W-2 and two field isolates of Plasmodium falciparum using lactate dehydrogenase assay. The products were also tested for their cytotoxicity on LLC/MK2 monkey kidney cells. The EtOAc extract exhibited a significant antiplasmodial activity (IC₅₀ = 11.15 μg/mL on W-2; 3.91 and 4.74 μg/mL on field CAM10 and SHF4 isolates, respectively), whereas the n-hexane fraction showed a weak activity (IC₅₀ = 73.78 μg/mL on W-2 and 21.85 μg/mL on SHF4). Three out of the four compounds showed good activity against all the three different parasite strains (IC₅₀ < 5 μM). Specicoside exhibited the highest activity on W-2 (IC₅₀ = 1.54 μM) followed by 2β, 3β, 19α-trihydroxy-urs-12-en-28-oic acid (IC₅₀ = 1.60 μM) and atranorin (IC₅₀ = 4.41 μM), while p-hydroxycinnamic acid was the least active (IC₅₀ = 53.84 μM). The EtOAc extract and its isolated compounds (specicoside and p-hydroxycinnamic acid) were non-cytotoxic (CC₅₀ > 30 μg/mL), whereas the n-hexane extract and two of its products, atranorin and 2β, 3β, 19α-trihydroxy-urs-12-en-28-oic acid showed cytotoxicity at high concentrations, with the last one being the most toxic (CC₅₀ = 9.37 μg/mL). These findings justify the use of K. africana stem bark as antimalaria by traditional healers of Western Cameroon, and could constitute a good basis for further studies towards development of new leads or natural drugs for malaria.