Biotools: an R function to predict spatial gene diversity via an individual-based approach.

The gene diversity or expected heterozygosity (HE) is based on the allele frequency and is often used as a measure of genetic variability of populations. Knowing the pattern of spatial distribution of HE can be useful for determining strategies of conservation and sampling of collections of individuals. In addition, it can allow one to detect genetic boundaries in a landscape. We adapted a Wombling method based on assignment tests in a circular moving window extensively sampled over the study area in order to estimate HE at points of a prediction grid. The function sHe(), package biotools, is an easy-to-use and flexible implementation in R language that accepts as input geographical and genotyping data. The package biotools is distribution-free under the GPL-2/3 license and currently available from the Comprehensive R Archive Network (CRAN) at . The R platform and all R dependencies are similarly available from CRAN.

Genetic diversity of arboreal cotton populations of the Brazilian semiarid: a remnant primary gene pool for cotton cultivars.

Mocó cotton belong to the same species as the cultivated species, Gossypium hirsutum, and cultivated forms were mainly landraces but also developed as cultivars, bearing good fiber quality and drought tolerant when cropped as a perennial species. The northeast Brazil crop system based on this cotton type is finished, with a few small area planted in the three main States, where it was previously cultivated (Ceará, Paraíba, and Rio Grande do Norte), but in others, maintenance is accomplished by single dooryard plants. Plants were found in all visited Northeast Brazil municipalities, sometimes in the North of the country, and were collected for ex situ preservation and evaluation. Most of seeds had no fuzz (62.2%) and 94.6% of the genotypes presented spot in flowers. Seventy-one alleles were revealed in 12 loci. The genetic structure of the population evaluated by microsatellite markers shows two main groups, one comprising the Seridó region where landraces were originated and other comprising the state of Ceará, where a specific breeding program was developed. Genotypes collected in North Brazil States as well as those collected in Bahia, Alagoas, and Sergipe grouped with those collected in Ceará. The Mantel correlogram indicates a significant (P < 0.05) correlation between genetic and geographical distances up to 77 km. The ex situ maintenance and agronomical evaluation are the main concerns for mocó, as the use of the agricultural interesting traits, possibly introgressed to other genotypes, is predicted. The in situ preservation is still of interest since there is more diversity there than in the collected plants and some should be continued due to use as medicinal plant.

Genetic diversity of high performance cultivars of upland and irrigated Brazilian rice.

The objective of this study was to analyze the diversity and discrimination of high-performance Brazilian rice cultivars using microsatellite markers. Twenty-nine rice cultivars belonging to EMBRAPA Arroz e Feijão germplasm bank in Brazil were genotyped by 24 SSR markers to establish their structure and genetic discrimination. It was demonstrated that the analyzed germplasm of rice presents an expressive and significant genetic diversity with low heterogeneity among the cultivars. All 29 cultivars were differentiated genetically, and were organized into two groups related to their upland and irrigated cultivation systems. These groups showed a high genetic differentiation, with greater diversity within the group that includes the cultivars for irrigated system. The genotyping data of these cultivars, with the morphological e phenotypical data, are valuable information to be used by rice breeding programs to develop new improved cultivars.

Implications of SNP weighting on single-step genomic predictions for different reference population sizes.

We investigated the importance of SNP weighting in populations with 2,000 to 25,000 genotyped animals. Populations were simulated with two effective sizes (20 or 100) and three numbers of QTL (10, 50 or 500). Pedigree information was available for six generations; phenotypes were recorded for the four middle generations. Animals from the last three generations were genotyped for 45,000 SNP. Single-step genomic BLUP (ssGBLUP) and weighted ssGBLUP (WssGBLUP) were used to estimate genomic EBV using a genomic relationship matrix (G). The WssGBLUP performed better in small genotyped populations; however, any advantage for WssGBLUP was reduced or eliminated when more animals were genotyped. WssGBLUP had greater resolution for genome-wide association (GWA) as did increasing the number of genotyped animals. For few QTL, accuracy was greater for WssGBLUP than ssGBLUP; however, for many QTL, accuracy was the same for both methods. The largest genotyped set was used to assess the dimensionality of genomic information (number of effective SNP). The number of effective SNP was considerably less in weighted G than in unweighted G. Once the number of independent SNP is well represented in the genotyped population, the impact of SNP weighting becomes less important.

Intraspecific differentiation of Hancornia speciosa revealed by simple sequence repeat and random amplified polymorphic DNA markers.

Hancornia speciosa, popularly known as mangabeira, is a fruit tree native to the Brazilian Cerrado that shows great economic potential, due to its multiple uses. Intraspecific classification of this species is difficult because it shows high morphological diversity. An early study of the species reported that there are six botanic varieties that differ morphologically mainly in the shapes of their leaves and flowers. Except to note the wide morphological variation and economic potential of this species, few studies have been published about the genetic diversity of mangabeira. Knowledge of the genetic variability of this species among populations would be useful for genetic conservation and breeding programs. Therefore, we tested the transferability of 12 simple sequence repeats from expressed sequence tags (EST-SSRs) from Catharanthus roseus to H. speciosa and used 10 random amplified polymorphic DNA markers to evaluate the genetic variability among botanical varieties of H. speciosa. We obtained a high transferability frequency of EST-SSR markers from C. roseus to H. speciosa (75%). However, EST-SSR markers showed low heterozygosity and locus variability (two or three alleles by locus), which suggest low genetic diversity in the mangabeira samples. The Jaccard dissimilarity index and an examination of geographic distances indicated a non-spatial structuring of the genetic variability. Our markers were unable to distinguish H. speciosa botanical varieties.

Natural hybridization between Gossypium mustelinum and exotic allotetraploid cotton species.

Cotton has been collected in Brazil for decades for its conservation, evaluation, and the use of its genetic resources. Gossypium mustelinum is an allotetraploid cotton species that only occurs in Brazil, and little is known about its genetic potential for improvement. However, the species is threatened by habitat fragmentation and interspecific hybridization with exotic species of cotton. In this study, we investigated the rate of natural hybridization in two populations of G. mustelinum in Bahia, Brazil, with G. hirsutum and G. barbadense using a set of microsatellite markers.

Discrimination of common bean cultivars using multiplexed microsatellite markers.

Analysis of DNA polymorphisms allows for the genetic identification and precise discrimination of species with a narrow genetic base such as common bean. The primary objectives of the present study were to molecularly characterize commercial common bean varieties developed at various research institutions using microsatellite markers and to determine the degree of genetic diversity among the bean varieties analyzed. Fifty cultivars representing 12 grain classes and 64 genitors, i.e., accessions used to develop these cultivars, were characterized. Based on an analysis of 24 simple sequence repeats, the estimates for the average number of alleles and genetic diversity were 8.29 and 0.646, respectively. The combined probability of identity was estimated at 7.05 x 10(-17), indicating a high individual discriminatory power. Thirty-two percent of the cultivars exhibited heterogeneity for multiple loci that reflected either homozygosity for different alleles of a given locus in different individuals or heterozygosity for the locus. The average genetic diversity for the groups of cultivars and genitors was 0.605 and 0.660, respectively, with no genetic differentiation (FST) between these groups. Although similar estimates of expected heterozygosity were observed when the cultivars were grouped by release date, a greater number of private alleles was observed in the most recent cultivars. The genetic differentiation among cultivars originating from different institutions was not different from zero (FST = 0.01). The molecular profile database derived from these analyses may increase the statistical power of genetic estimates and may be incorporated into breeding programs for common bean. Furthermore, the profiles obtained for the different cultivars may be used as molecular descriptors to complement traditional descriptors used in distinctiveness, uniformity and stability tests, thereby improving the traceability of samples and their derivatives and helping to protect the intellectual property rights of breeders.

Antimicrobial effect of Lippia sidoides and thymol on Enterococcus faecalis biofilm of the bacterium isolated from root canals.

The species Lippia sidoides Cham. (Verbenaceae) is utilized in popular medicine as a local antiseptic on the skin and mucosal tissues. Enterococcus faecalis is the bacterium isolated from root canals of teeth with persistent periapical lesions and has the ability to form biofilm, where it is responsible for the failure of endodontic treatments. Essential oil of L. sidoides (EOLS) and its major component, thymol, were evaluated for reducing the CFU in biofilms of E. faecalis in vitro. The essential oil was obtained by hydrodistillation and examined with respect to the chemical composition, by gas chromatography-mass spectrometry (GC-MS). The GC-MS analysis has led to the identification of thymol (84.9%) and p-cymene (5.33%). EOLS and thymol reduced CFU in biofilms of E. faecalis in vitro (time of maturation, 72 h), with an exposure time of 30 and 60 min at concentrations of 2.5 and 10%. There was no statistical difference in effect between EOLS and thymol, demontrating that this phenolic monoterpene was the possible compound responsible for the antimicrobial activity of EOLS. This study provides a basis for the possible utilization of EOLS as an adjuvant in the treatment of root canals that show colonization by E. faecalis.

Bioaccessibility and bioavailability of essential and potentially toxic trace elements in potato cultivars: A comprehensive nutritional evaluation.

Among the most consumed foods in the world is potato, which occupies the first place as a non-grain commodity, demonstrating the importance of its assessment concerning the population's food safety. In this study, the nutrients Ca, Mg, K, P, Cu, Mn, Fe, and Zn and the potentially toxic trace elements Cd, Cr, and Pb were evaluated considering their total contents, bioaccessible and bioavailable fractions in different potato cultivars, in an unpublished approach in the literature. The in vitro standard gastrointestinal digestion method (INFOGEST) and a model of the intestinal epithelial barrier using the Caco-2 cell line were applied for investigate the presence of metals in potato. For the macroelements, the bioaccessibility (% w/w) varied in the ranges: K (57-72 %), P (59-76 %), Mg (83-103 %), and Ca (30-123 %), whereas for the microelements were: Cu (27-74 %) and Mn (4.22-12.02, 60-119 %). The potentially of trace toxic elements, Cd and Pb, were found in 75 % of the samples, however, all the concentration values were below the maximum levels allowed of 0.10 µg/g. Chromium was determined only in potato peels and has no maximum established level. The bioaccessible and bioavailable fractions of Cd, Cr, and Pb were below the limits of quantification of the spectrometric methods (LOQ - µg/L: 0.063 Cd, 0.65 Cr, and 0.44 Pb). The potato samples were considered safe for consumption regarding the presence of potentially toxic trace elements, with a remarkable nutritional contribution.

Molecular characterization of high performance inbred lines of Brazilian common beans.

The identification of germplasm genetic variability in breeding programs of the common bean (Phaseolus vulgaris) is essential for determining the potential of each combination of parent plants to obtain superior genotypes. The present study aimed to estimated the extent of genetic diversity in 172 lineages and cultivars of the common bean by integrating five tests of value for cultivation and use (VCU) that were conducted over the last eight years by the breeding program of Embrapa Arroz e Feijão in Brazil. Nine multilocus genotyping systems composed of 36 fluorescent microsatellite markers distributed across 11 different chromosomes of the common bean were used, of which 24 were polymorphic in all trials. One hundred and eighty-seven alleles were identified, with an average of 7.79 alleles per locus and an average gene diversity of 0.65. The combined probability of identity for all loci was 1.32 x 10(-16). Lineages that are more genetically divergent between the selection cycles were identified, allowing the breeding program to develop a crossbreed between elite genotypes with a low degree of genetic relatedness. HE values ranged from 0.31 to 0.63, with a large reduction in the genetic base over successive selection cycles. The test showed a significant degree of differentiation (FST = 0.159). Private alleles (26%) were identified and can be directly incorporated into the gene pool of cultivated germplasm, thereby contributing effectively to the expansion of genetic diversity in this bean-breeding program.

Antinociceptive and anti-inflammatory activities of Hymenaea martiana Hayne (Fabaceae) in mice.

Hymenaea martiana is a species popularly known in Northeastern Brazil as "jatobá" and used in folk medicine to treat pain and inflammation. The aim of this work was to evaluate the antinociceptive and anti-inflammatory activity of H. martiana. In the present study, we carried out an investigation about the effects of the crude ethanolic extract (Hm-EtOH) and the ethyl acetate fraction (Hm-AcOEt) in models of nociception and inflammation in mice. Chemical (acetic acid-induced writhing and formalin) and thermal stimuli (hot plate) were used for the evaluation of antinociceptive activity, while for the anti-inflammatory profile paw edema induced by carrageenan was used, along with leukocyte migration to the peritoneal cavity. The presence of the flavonoid astilbin in the samples was characterized through HPLC-DAD-MS analysis. Hm-EtOH and Hm-AcOEt (100, 200 and 400 mg.kg-1, i.p.) significantly reduced the number of abdominal contortions and decreased the paw licking time in the formalin test. In the hot plate, the extract increased the latency time of animals. Hm-EtOH and Hm-AcOEt inhibited significantly the increase in the edema after the administration of carrageenan. Hm-EtOH and Hm-AcOEt inhibited leukocyte migration in the peritonitis test. HPLC-DAD-MS analysis of Hm-EtOH and Hm-AcOEt revealed the presence of the flavonoid astilbin in the samples. According to the results of this study, both Hm-EtOH and Hm-AcOEt have antinociceptive and anti-inflammatory activities, which could be related with the presence of flavonoid in the extracts. The results reinforce the popular use of this plant.

Gastroprotective and cicatrizing activity of the Ziziphus joazeiro Mart. leaf hydroalcoholic extract.

Ziziphus joazeiro Mart., popularly known as 'juazeiro', is a species used in popular medicine for the treatment of bronchitis, gastric ulcers, skin wounds, and in the manufacture of cosmetic and food products. The objective of this study is to evaluate the gastroprotective and cicatrizing activity of the Z. joazeiro Mart. leaf hydroalcoholic extract (EHFZJ). The acute pre-clinical toxicity was determined by the single administration of the EHFZJ (2000 mg/kg/p.o.) and by assessing clinical signs of toxicity, according to established criteria by Malone, or mortality. Gastroprotective activity was identified through classical models of acute gastric lesions induced by indomethacin, absolute and acidified ethanol (100, 200 and 400 mg/kg/per os) and the physical barrier mechanism (400 mg/kg/per os or intraperitoneally). The cicatrizing activity of the EHFZJ was investigated by measuring the speed of wound closure and the percentage of contraction. The acute pre-clinical toxicity of EHFZJ showed no signs of toxicity and mortality. The EHFZJ demonstrated a gastroprotective effect at the 400 mg/kg dose in the classical models of acute gastric injury induced by indomethacin, absolute and acidified ethanol. The EHFZJ administration (orally) demonstrated significant inhibition, suggesting a possible physical barrier mechanism exists. The EHFZJ showed no significant differences in terms of percentage of contraction or the speed of wound closure during the observation times (0, 3, 7, 11 and 14 days). The results obtained in this study provide evidence of a potential gastroprotective activity for the Ziziphus joazeiro Mart. Leaf hydroalcoholic extract.

Risk factors for overweight and obesity in adolescents of a Brazilian University: a case-control study.

OBJECTIVE: To analyze the risk factors associated to overweight and obesity in freshmen of a public university in the Center-West region of Brazil. METHODS: A case-control study comprising 1,465 adolescents, identifying 106 cases (Body Mass Index (BMI)>or = 85th percentile of National Center for Health Statistics -NCHS) and 233 controls (BMI > 5th and < 85th percentile of NCHS). Interviews were made to collect information on socioeconomic data, eating habits, physical activity and health habits. Information on parents' height and weight were obtained over the telephone. RESULTS: Non-conditional multivariate and hierarchical logistic regression showed that overweight and obesity were positively associated to males (odds ratio (OR) 2.25, 95% confidence interval (CI) 1.37-3.69), mother's BMI (OR 3.45, 95% CI 2.16-5.8), overweight in childhood (OR 2.81, 95% CI 1.62-4.85), consumption of fruits less than once a day (OR 1.84, 95% CI 1.05-3.21) and the habit of weight-loss dieting (OR 6.33, 95% CI 2.68-14.94). CONCLUSION: Results emphasize the need for educational interventions at early ages involving the whole family to control the excess of weight.

Drought in Portugal: Current regime, comparison of indices and impacts on extreme wildfires.

In Portugal, drought characterizes the climatic variability, contributes to the increase of fire risk and its duration and intensity are expected to increase in future climate. Surprisingly, the quantitative and objective analysis to characterize the drought regime in current climate conditions as well as its influence on the occurrence of large wildfires (LW) has never been done for Portugal, which are the main objectives of this study. We assessed drought regime for recent past climate conditions (1981-2017), using four different drought indices, namely SPI, SPEI, RDI and VCI, and assessed the influence of drought in LW occurrence. Results include the characterization of drought number, duration, severity, intensity, extension, intra- and inter-annual variability for different classes of severity and the space-time distribution of LW in drought periods and affected area. Our main findings include 67% of the study period were drought months; regions with higher drought duration and severity assessed with SPI and SPEI for general drought conditions evolves from north to south with the increase of drought assessment period; drought characteristics present low intra - annual and inter - annual variability but are clearly associated to the temporal and spatial distribution of LW. In fact, all LW occurred during drought assessed with SPI or SPEI, almost all LW (97% to 95%) and corresponding burnt area (98% to 97%) occurred during drought assessed with SPI and SPEI. The relationship between drought and fire incidence is statistical significant for 3 - month SPI, 3 - and 6 - month SPEI, and is particularly strong for Moderate and Severe drought. 85% and 87% of LW occurred in area affected by drought assessed with SPI or SPEI, respectively. It is not clear which is the best index, but drought plays a fundamental role in the occurrence of large wildfires in Portugal.

The antioxidative effects of bioactive products from Sargassum polycystum C. Agardh and Sargassum duplicatum J. Agardh against inflammation and other pathological issues.

The present study was aimed to determine the phenol, total flavonoids and antioxidant potentials of Sargassum polycystum C. Agardh and Sargassum duplicatum J. Agardh from south east coast of Tamil Nadu and India using DPPH, phophomolybdenum and hydrogen peroxide scavenging activity. The total phenols, total flavonoids and antioxidant activities of S. polycystum and S. duplicatum were determined. Highest phenols (33.49 and 149.52 mg GAE/g) were observed in chloroform extracts of S. polycystum and methanolic extracts of S. duplicatum. The acetone extracts of S. polycystumand S. duplicatum demonstrated maximum amount of flavonoids compared to other studied extracts. Maximum amount of phosphomolybdenum reduction was observed in acetone extracts of S. polycystum and methanolic extracts of S. duplicatum. The DPPH radical scavenging activity of different extracts of S. polycystum and S. duplicatum were as follows acetone > chloroform > Petroleum ether > methanol and acetone > chloroform > methanol >Petroleum ether respectively. The H2O2 scavenging activity of S. polycystum extracts were as follows Petroleum ether extracts of S. polycystum (67.9%) > acetone (67.3%) > chloroform (58.6%) > methanol (51.78%). Acetone extracts of S. duplicatum showed maximum inhibition (90.39%) followed by petroleum ether, chloroform and methanolic extracts (75.11, 72.37 and 54.59%) respectively. The present study results confirmed the antioxidant properties of the two selected brown seaweeds viz., S. polycytum and S. duplicatum. The total phenols, flavonoids and alkaloids may be responsible for the antioxidant activities.

Larvicidal activity of some medicinal plant extracts against filariasis fever mosquito, Culex quinquefasciatus (Say.) (Diptera: Culicidae).

OBJECTIVES: The present study was aimed to find the larvicidal activity of petroleum ether, chloroform, acetone and methanolic extracts of Dichanthium foveolatum (Del.) Roberty, Leptochloa uniflora Hochst, Pancratium triflorum Roxb and Molineria trichocarpa (Wight) N.P.Balakr against Culex quinquefasciatus. METHODS: The larvicidal potential of selected plant extracts were determined against 4th instar larvae of C. quinquefasciatus with various concentrations viz., 50, 100, 150, 200 and 250 mg/ml. The mortality counts were made after 24 h of incubation and LC50 values were calculated. RESULTS: Chloroform extracts of studied plants were showed highest larvicidal activity with remarkable irritant against the larva of C. quinquefasciatus. Highest larvicidal activity was observed in the chloroform extract of D. foveolatum against the larva of C. quinquefasciatus with LC50 = 277.03 mg/ml. The larvicidal activity of the studied plants as follows chloroform extract of D. foveolatum (LC50 = 277.03 mg/ml) >L. uniflora (LC50 = 300.56 mg/ml) >M. trichocarpa (LC50 = 306.60 mg/ml) >P. triflorum (LC50 318.42 mg/ml). The larvicidal potential of P. triflorum was as follows Chloroform > acetone > methanol > petroleum ether. The larvicidal activities of L. uniflora and M. trichocarpa were as follows Chloroform > petroleum ether > acetone > methanol respectively. The larvicidal potentials of D. foveolatum was as follows Chloroform > methanol > acetone > petroleum ether. CONCLUSION: The chloroform extract of D. foveolatum find use as broad-spectrum larvicidal agent in the near future. It is hoped that more work would be undertaken to evaluate the utility of these plant extracts for field applications considering the promising leads given by the present study.

Pancratium triflorum Roxb. (Amaryllidaceae) and Molineria trichocarpa (Wight) N.P. Balakr (Hypoxidaceae): Cytotoxic and antioxidant activities.

The present investigation was carried out to evaluate the antioxidant and cytotoxic activities of Pancratium triflorum Roxb and Molineria trichocarpa were collected from South Vagaikulam, Tirunelveli district, Tamil Nadu, India. The antioxidant activities of P. triflorum extracts were as follows with the IC50 values methanol (228.13 µg/mL) > chloroform (311.33 µg/mL) > acetone (398.08 µg/mL) > petroleum ether (410.16 µg/mL). The antioxidant activities of P. triflorum and M. trichocarpa extracts were as follows with the IC50 values methanol (80.93 µg/mL) > acetone (98.02 µg/mL) > chloroform (186.84 µg/mL) > petroleum ether (209.64 µg/mL). Among the various extracts of P. triflorum, methanolic extracts showed the strongest phosphomolybdenum reduction (140.56 g AA/100 g). Among the tested extracts, acetone extracts of M. trichocarpa showed maximum inhibition with 71.36 ±â€¯5.86%. In P. triflorum, chloroform extracts showed maximum inhibition (69.51%). The petroleum ether extract of M. trichocarpa was found to be most effective at which 50% mortality (LC50) and 90% mortality (LC90) of brine shrimp nauplii were found to be 29.22 and 184.82 mg/mL. This study results revealed the antioxidant and cytotoxic properties of P. triflorum and M. trichocarpa. Further investigations are needed to isolate and validate the active principles of the extract responsible various pharmacological properties.

Vasorelaxant effect of the Lippia alba essential oil and its major constituent, citral, on the contractility of isolated rat aorta.

The Lippia alba (Mill.) N.E. Brown (Verbenaceae) species popularly known as lemon balm has sedative, analgesic and spasmolytic properties. This study aimed to evaluate the vasorelaxant effect of the L. alba essential oil (EOLa) and its major constituent, citral, rat on aorta. Isometric muscle contraction were induced by potassium (K 60 mM) or phenylephrine (PHE, 0.1 µM) in isolated aortic rings. EOLa and citral promoted a smooth muscle relaxant action, which was potentiated by the presence of the endothelium; PHE-induced contractions (0.1 µM) in aorta with endothelium, had EC50 values of 352.73 ± 19.39 µg/mL and 99.34 ± 7.2 µg/mL for EOLa and citral, respectively. In the presence of a nitric oxide synthase inhibitor, L-NAME, the EC50 values were 654.19 ± 10.46 µg/mL and 601.66 ± 10.922 µg/mL for EOLa and citral, respectively. EOLa and citral dose-dependently relaxed contractions induced by BAY-K 8644, a calcium channel agonist, and by Phorbol 12,13-dibutyrate an activator of protein kinase C. EOLa and citral produced a vasorelaxant effect in isolated aorta which was potentiated by the presence of endothelium. In summary, EOLa and citral, probably using several mechanisms of action, relaxed aortic smooth muscle with maximal pharmacologic efficacy.

Modulation of antibiotic activity by the hydroalcoholic extract from leaves of Caryocar coriaceum WITTM.

The aim of this study was to identify the presence of tannins, phenols and flavonoids on the hydroalcoholic extract of Caryocar coriaceum leaves (HECCL) and to determine the antioxidant and antibacterial activity of this extract. The extract was tested alone (1024-1 µg/mL) or associated (MIC/8) with several antibiotics in order to identify any antibacterial activity against multiresistant bacterial strains (Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa). The existence of tannins, total phenols (901.31 mg/g) and flavonoids (89.68 mg/g) was confirmed in the HECCL. The presence of rutin and quercetin were confirmed by Thin-layer chromatography (TLC). Using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) method, the antioxidant activity of the extract (9 µg/mL) was determined. Moreover, the Minimum Inhibitory Concentration (MIC) value found for HECCL was 1024 µg/mL and the association between HECCL (MIC/8) with benzylpenicillin significantly changed its minimum inhibitory concentration from 2500 to 625 µg/mL against E. coli.