Determination and quantification of sinigrin glucosinolates in Alternaria solani susceptible tomato (Solanum lycopersicum) leaves treated with moringa (Moringa oleifera) leaf extract

Abstract

The study investigates the presence and quantity of antimicrobial sinigrin glucosinolates in tomato leaves after spraying them with moringa (Moringa oleifera) leaf extract (MLAE). Moringa concentrates (0.5, 0.75, 1.00 and 1.5?kg?L^?1 (w v^?1)) were prepared. Distilled water was the control. Sampled tomato leaves were air-dried, freeze-dried and extracted firstly using pure methanol in a hot water bath and then pellet re-extracted using 5?mL of hot aqueous methanol (70% v v^?1). An ion exchange column, and sulphatase was used to achieve glucosiodesulphonation. High performance liquid chromatography (HPLC) was employed in the identification and quantitative analysis of the sinigrin glucosinolates. Tomato (Solanum lycopersicum) leaves treated with MLAE revealed highly significant (p?<?.001) content of sinigrin glucosinolates. The sinigrin standard and the desulphated sinigrin glucosinolates had a 7?s retention time difference; 5?kg?L^?1 (w v^?1) resulted in a superior amount of sinigrin in tomato leaves as compared to all the other MLAE concentrations. The study reveals that spraying MLAE on putatively diseased tomato leaves donates specific quantifiable glucosinolates like sinigrin, which may be involved in defense against tomato diseases and, hence, recommends use of 5?kg?L^?1 (w v^?1) for the highest sinigrin defense tag.     

First experimental evidence of corals feeding on seagrass matter

We present the first experimental evidence of a coral (Oulastrea crispata) ingesting and assimilating seagrass material. Tropical seagrass meadows export a substantial portion of their productivity and can provide an important source of nutrients to neighbouring systems such as coral reefs; however, little is known about the mechanisms of this link. To investigate whether seagrass nutrient uptake via coral heterotrophy is possible, we conducted a feeding experiment with seagrass particulate and dissolved organic matter. Using gut extractions and stable isotope analyses, we determined that O. crispata ingested ¹⁵N-enriched seagrass particles and assimilated the nitrogen into its tissue at a rate of 0.75 μg N cm^?2 h^?1. Corals took up nitrogen from dissolved matter at a comparable rate of 0.98 μg N cm^?2 h^?1. While other ecological connections between seagrass meadows and reef ecosystems are well known, our results suggest a previously unstudied direct nutritional link between seagrasses and corals.     

Lipoxygenase6-Dependent Oxylipin Synthesis in Roots Is Required for Abiotic and Biotic Stress Resistance of Arabidopsis

Jasmonates are oxylipin signals that play important roles in the development of fertile flowers and in defense against pathogens and herbivores in leaves. The aim of this work was to understand the synthesis and function of jasmonates in roots. Grafting experiments with a jasmonate-deficient mutant demonstrated that roots produce jasmonates independently of leaves, despite low expression of biosynthetic enzymes. Levels of 12-oxo-phytodienoic acid, jasmonic acid, and its isoleucine derivative increased in roots upon osmotic and drought stress. Wounding resulted in a decrease of preformed 12-oxo-phytodienoic acid concomitant with an increase of jasmonic acid and jasmonoyl-isoleucine. 13-Lipoxygenases catalyze the first step of lipid oxidation leading to jasmonate production. Analysis of 13-lipoxygenase-deficient mutant lines showed that only one of the four 13-lipoxygenases, LOX6, is responsible and essential for stress-induced jasmonate accumulation in roots. In addition, LOX6 was required for production of basal 12-oxo-phytodienoic acid in leaves and roots. Loss-of-function mutants of LOX6 were more attractive to a detritivorous crustacean and more sensitive to drought, indicating that LOX6-derived oxylipins are important for the responses to abiotic and biotic factors.Oxylipins are ubiquitous signaling molecules that are derived from polyunsaturated fatty acids by enzymatic and nonenzymatic processes. In plants, the biosynthesis and function of oxylipins of the jasmonate family in aboveground tissues has been investigated in detail. Jasmonates comprise 12-oxo-phytodienoic acid (OPDA), jasmonic acid (JA), and derivatives of JA. In leaves, jasmonates accumulate in response to abiotic factors such as wounding, drought, osmotic stress, darkness, and ozone and during interactions with organisms such as herbivores, pathogens, and mutualistic organisms (Wasternack, 2007). The relevance of jasmonates in wound response, ozone tolerance, and the defense against herbivores and necrotrophic pathogens in leaves has been well investigated using mutants in JA biosynthesis and signaling (Browse, 2009a). In addition, jasmonates play an important role in flower development, and Arabidopsis (Arabidopsis thaliana) mutants in the JA pathway are male sterile (Browse, 2009b). The first step in jasmonate biosynthesis is catalyzed by 13-lipoxygenases (LOXs). The resulting 13(S)-hydroperoxyoctadecatrienoic acid (13-HPOTE) is converted by allene oxide synthase (AOS) and allene oxide cyclase to OPDA (Wasternack, 2007). These enzymatic steps are located in plastids. OPDA is transported to peroxisomes and converted to JA. JA can be further metabolized to different derivatives that take place mainly in the cytosol. The conjugation of JA with Ile is an important step because jasmonoyl-Ile (JA-Ile) has been identified as a biologically active jasmonate (Staswick and Tiryaki, 2004). OPDA is also biologically active without conversion to JA derivatives. In contrast to all other jasmonates, the OPDA structure contains an electrophilic α,β-unsaturated carbonyl group that renders OPDA more reactive than JA. Therefore, OPDA is classified as a reactive electrophile species with unique signaling properties different from other jasmonates (Farmer and Davoine, 2007).Of the six lipoxygenase genes present in Arabidopsis, four genes encode 13-LOX. For the respective enzymes LOX2, LOX3, LOX4, and LOX6, it was shown that linolenic acid is the preferred substrate and that 13-HPOTE is formed in vitro (Bannenberg et al., 2009). All four enzymes are proposed to be located in plastids. LOX2 is highly expressed in leaves; expression is up-regulated by jasmonates and stress treatments such as wounding and osmotic stress (Bell and Mullet, 1993; Seltmann et al., 2010a). LOX2 was shown to contribute the majority of jasmonate synthesis upon wounding and osmotic stress and during senescence in leaves (Bell et al., 1995; Glauser et al., 2009). LOX2 is also responsible for the accumulation of arabidopsides (Glauser et al., 2009), which are galactolipids containing esterified OPDA in plastids by direct oxidation of galactolipids (Zoeller et al., 2012). LOX3 and LOX4 are required for the development of fertile flowers (Caldelari et al., 2011). LOX6 shows overall low expression (Bannenberg et al., 2009). Recently, it was reported that LOX6 contributes to the fast accumulation of JA and JA-Ile in wounded leaves and is required for the fast increase of JA and JA-Ile in distal leaves after wounding (Chauvin et al., 2013).In contrast to leaves and flowers, little is known on jasmonate biosynthesis and function in roots. Expression of the plastid-localized enzymes of jasmonate synthesis LOX2, AOS, and allene oxide cyclase2 is very low in roots (Zimmermann et al., 2004). By contrast, enzymes such as 9-LOX and α-dioxygenase1 are strongly expressed in roots. These enzymes are involved in the biosynthesis of oxylipins different from jasmonates, and 9-LOX products have been shown to regulate lateral root development because mutants in LOX1 and LOX5 produce more lateral roots (Vellosillo et al., 2007). However, jasmonate function in roots is still obscure. Here, we analyzed jasmonate accumulation in roots upon different stress treatments and show that mutants defective in LOX6 are impaired in stress-induced jasmonate synthesis and are more susceptible to drought and detritivore feeding.     

Patterns of Infection with Cryptosporidium sp. and Giardia sp. in Three Species of Free-Ranging Primates in the Peruvian Amazon

Recent evidence of pathogen transmission to humans from wild primates and a greater recognition of the risk of human pathogen transmission to free-ranging primates have raised awareness of the potential impact of zoonotic pathogen transmission on primate conservation and nonhuman primate and human health. Cryptosporidium and Giardia are zoonotic protozoan parasites transmitted via fecal–oral contamination or water that can cause gastritis or enteritis in human and nonhuman primates. From June 2002 to September 2003, we collected fecal samples noninvasively from two species of tamarins (Saguinus mystax and S. nigrifrons) and one species of titi monkeys (Callicebus cupreus) at the Estación Biológica Quebrada Blanco in the Peruvian Amazon to determine the distribution and prevalence of these potential pathogens. We screened 140 fecal samples representing known individuals of each species for Cryptosporidium and Giardia using the Merifluor immunoflourescence assay to determine the prevalence and intensity of infection with these organisms. With the exception of two samples we collected during the same week from a juvenile male Saguinus mystax, all samples were negative for Cryptosporidium. None of the fecal samples were positive for Giardia. The low prevalence of infection we observed limited our ability to examine the effects of demographic and environmental variables on patterns of infection; however, the exceptionally low prevalence of Cryptosporidium suggests that it is not a current health threat to these primate populations. Although the origin of infection with Cryptosporidium in the juvenile male Saguinus mystax cannot be determined, its presence alerts us to the potential for cross-species transmission and highlights the need for more detailed research to improve our understanding of the distribution and diversity of potentially pathogenic protozoa in Neotropical primate populations.     

NADP-Specific Electron-Bifurcating [FeFe]-Hydrogenase in a Functional Complex with Formate Dehydrogenase in Clostridium autoethanogenum Grown on CO

Flavin-based electron bifurcation is a recently discovered mechanism of coupling endergonic to exergonic redox reactions in the cytoplasm of anaerobic bacteria and archaea. Among the five electron-bifurcating enzyme complexes characterized to date, one is a heteromeric ferredoxin- and NAD-dependent [FeFe]-hydrogenase. We report here a novel electron-bifurcating [FeFe]-hydrogenase that is NADP rather than NAD specific and forms a complex with a formate dehydrogenase. The complex was found in high concentrations (6% of the cytoplasmic proteins) in the acetogenic Clostridium autoethanogenum autotrophically grown on CO, which was fermented to acetate, ethanol, and 2,3-butanediol. The purified complex was composed of seven different subunits. As predicted from the sequence of the encoding clustered genes (fdhA/hytA-E) and from chemical analyses, the 78.8-kDa subunit (FdhA) is a selenocysteine- and tungsten-containing formate dehydrogenase, the 65.5-kDa subunit (HytB) is an iron-sulfur flavin mononucleotide protein harboring the NADP binding site, the 51.4-kDa subunit (HytA) is the [FeFe]-hydrogenase proper, and the 18.1-kDa (HytC), 28.6-kDa (HytD), 19.9-kDa (HytE1), and 20.1-kDa (HytE2) subunits are iron-sulfur proteins. The complex catalyzed both the reversible coupled reduction of ferredoxin and NADP⁺ with H₂ or formate and the reversible formation of H₂ and CO₂ from formate. We propose the complex to have two functions in vivo, namely, to normally catalyze CO₂ reduction to formate with NADPH and reduced ferredoxin in the Wood-Ljungdahl pathway and to catalyze H₂ formation from NADPH and reduced ferredoxin when these redox mediators get too reduced during unbalanced growth of C. autoethanogenum on CO (E_0′ = −520 mV).     

Immunologic evaluation and validation of methods using synthetic peptides derived from Mycobacterium tuberculosis for the diagnosis of tuberculosis infection

The goal of this study was to demonstrate the usefulness of an enzyme-linked immunosorbent assay (ELISA) for the serodiagnosis of pulmonary tuberculosis (PTB) and extrapulmonary TB (EPTB). This assay used 20 amino acid-long, non-overlapped synthetic peptides that spanned the complete Mycobacterium tuberculosis ESAT-6 and Ag85A sequences. The validation cohort consisted of 1,102 individuals who were grouped into the following five diagnostic groups: 455 patients with PTB, 60 patients with EPTB, 40 individuals with non-EPTB, 33 individuals with leprosy and 514 healthy controls. For the PTB group, two ESAT-6 peptides (12033 and 12034) had the highest sensitivity levels of 96.9% and 96.2%, respectively, and an Ag85A-peptide (29878) was the most specific (97.4%) in the PTB groups. For the EPTB group, two Ag85A peptides (11005 and 11006) were observed to have a sensitivity of 98.3% and an Ag85A-peptide (29878) was also the most specific (96.4%). When combinations of peptides were used, such as 12033 and 12034 or 11005 and 11006, 99.5% and 100% sensitivities in the PTB and EPTB groups were observed, respectively. In conclusion, for a cohort that consists entirely of individuals from Venezuela, a multi-antigen immunoassay using highly sensitive ESAT-6 and Ag85A peptides alone and in combination could be used to more rapidly diagnose PTB and EPTB infection.     

A population-based study of first and second-line drug-resistant tuberculosis in a high-burden area of the Mexico/United States border

The resistance of 139 Mycobacterium tuberculosis (MTB) isolates from the city of Monterrey, Northeast Mexico, to first and second-line anti-TB drugs was analysed. A total of 73 isolates were susceptible and 66 were resistant to anti-TB drugs. Monoresistance to streptomycin, isoniazid (INH) and ethambutol was observed in 29 cases. Resistance to INH was found in 52 cases and in 29 cases INH resistance was combined with resistance to two or three drugs. A total of 24 isolates were multidrug-resistant (MDR) resistant to at least INH and rifampicin and 11 MDR cases were resistant to five drugs. The proportion of MDR-TB among new TB cases in our target population was 0.72% (1/139 cases). The proportion of MDR-TB among previously treated cases was 25.18% (35/139 cases). The 13 polyresistant and 24 MDR isolates were assayed against the following seven second-line drugs: amikacin (AMK), kanamycin (KAN), capreomycin (CAP), clofazimine (CLF), ethionamide (ETH), ofloxacin (OFL) and cycloserine (CLS). Resistance to CLF, OFL or CLS was not observed. Resistance was detected to ETH (10.80%) and to AMK (2.70%), KAN (2.70%) and CAP (2.70%). One isolate of MDR with primary resistance was also resistant to three second-line drugs. Monterrey has a high prevalence of MDR-TB among previously treated cases and extensively drug-resistant-MTB strains may soon appear.     

Changing distributions of ticks: causes and consequences

Today, we are witnessing changes in the spatial distribution and abundance of many species, including ticks and their associated pathogens. Evidence that these changes are primarily due to climate change, habitat modifications, and the globalisation of human activities are accumulating. Changes in the distribution of ticks and their invasion into new regions can have numerous consequences including modifications in their ecological characteristics and those of endemic species, impacts on the dynamics of local host populations and the emergence of human and livestock disease. Here, we review the principal causes for distributional shifts in tick populations and their consequences in terms of the ecological attributes of the species in question (i.e. phenotypic and genetic responses), pathogen transmission and disease epidemiology. We also describe different methodological approaches currently used to assess and predict such changes and their consequences. We finish with a discussion of new research avenues to develop in order to improve our understanding of these host–vector–pathogen interactions in the context of a changing world.