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S. Lambert

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Biologist with PhD in Biochemistry from Pasteur Institute in Paris. Worked for 26 years with Mars, Inc. as Asia/Pacific cocoa research manager in Brazil, Indonesia, Vietnam, PNG and Philippines on cocoa Hybrid trials/Clonal selections, Integrated Pest Management and cocoa fermentation/quality. Retired in 2020 with some ongoing consultancy work.

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Papers by S. Lambert

Cementing proteins provide extra mechanical stabilization to viral cages

The study of virus shell stability is key not only for gaining insights into viral biological cyc... more The study of virus shell stability is key not only for gaining insights into viral biological cycles but also for using viral capsids in materials science. The strength of viral particles depends profoundly on their structural changes occurring during maturation, whose final step often requires the specific binding of 'decoration' proteins (such as gpD in bacteriophage lambda) to the viral shell. Here we characterize the mechanical stability of gpD-free and gpD-decorated bacteriophage lambda capsids. The incorporation of gpD into the lambda shell imparts a major mechanical reinforcement that resists punctual deformations. We further interrogate lambda particle stability with molecular fatigue experiments that resemble the sub-lethal Brownian collisions of virus shells with macromolecules in crowded environments. Decorated particles are especially robust against collisions of a few k B T (where k B is the Boltzmann's constant and T is the temperature B300 K), which approximate those anticipated from molecular insults in the environment.

Potential proteolytic activity of human plasma fibronectin: fibronectin gelatinase

by xavier blondeau and S. Lambert

Biological Chemistry Hoppe-Seyler, 1990

Human plasma fibronectin contains a latent proteinase that after activation cleaves gelatin and f... more Human plasma fibronectin contains a latent proteinase that after activation cleaves gelatin and fibronectin. The autoactivation propensity of the two purified cathepsin D-produced fragments of fibronectin (190 and 120 kDa) was compared. Both polypeptides were spontaneously activated in the presence of Ca2+. This activation was inhibited by EDTA. The active gelatinase was isolated from the autodigest of the 190-kDa fragment. Among various protein substrates, including laminin and native type I and IV collagens, the purified enzyme degraded only gelatin and fibronectin. We have named this proteinase FN-gelatinase. FN-gelatinase is inhibited by phenylmethanesulfonyl fluoride and also by pepstatin A like retroviral aspartic proteinases. The amino-acid composition of the purified enzyme (35 kDa) was compared with the entire fibronectin sequence using the computer programme FIT. The optimal fit indicated that the 35-kDa fragment corresponds to the stretch # 1043-1404. This sequence contai...

Learning lessons from participatory breeding research with Sulawesi cocoa smallholders

by S. Lambert and Agung Susilo

The similarity of cocoa flavour of fermented seeds from fingerprinted genotypes of Theobroma cacao L. from Brazil and Malaysia

Fermentation of Theobroma cacao L. seeds has been considered to be the most important factor infl... more Fermentation of Theobroma cacao L. seeds has been considered to be the most important factor influencing cocoa flavour and has accordingly received most attention. Development of a test procedure in which small amounts of wet cocoa seeds in net bags are inserted in fermentations of wet seeds from single or mixed genotypes has allowed the flavour potential of a wide range of genotypes to be examined. Flavour profiles of cocoa liquors from genotypes grown at BAL Plantations Sdn Bhd, Malaysia differed consistently for various replicate preparations indicating a strong genetic contribution to flavour. In the present study, similar flavour profiles were developed from cocoa from the same genotypes grown in Brazil and Malaysia and subjected to the same conditions of post-harvest processing. Genetic fingerprinting by Rapid Amplified Polymorphic DNA (RAPD) analysis confirmed that most of the genotypes tested for flavour in Brazil and Malaysia were the same, although there were also a few er...

[Proteolytic potential of fibronectin and degradation of extracellular matrix]

Pathologie-biologie, 1990

Fibronectin is one of the major adhesive glycoproteins and bears interaction sites for both cell ... more Fibronectin is one of the major adhesive glycoproteins and bears interaction sites for both cell receptors and the extracellular matrix. Disappearance of fibronectin is the first step of cellular transformation in carcinogenesis. This phenomenon has been ascribed to increased proteolysis of fibronectin or of its cellular receptor. Results obtained during previous studies by the authors have shown that the fibronectin molecule has latent proteolytic activities which become apparent only after the action of other external proteases. Two proteinases, FN-gelatinase and FN-laminase, were identified in cathepsin D fibronectin digest. The acute activity of these two proteases is responsible for degradation of the extracellular matrix. Furthermore, the sequences and functions of both enzymes share a number of features with retroviral proteases.

project Improving cocoa production through farmer involvement in demonstration trials of potentially superior and pest/disease resistant genotypes and integrated management practices

by Jeff Neilon and S. Lambert

Potential Proteolytic Activity of Fibronectin: Fibronectin Laminase and its Substrate Specificity

Biological Chemistry Hoppe-Seyler, 1990

The purified 190-kDa fibronectin fragment produced by cathepsin D can be spontaneously activated ... more The purified 190-kDa fibronectin fragment produced by cathepsin D can be spontaneously activated in the presence of CaCl2. This activation generates new proteolytic activities and also results in the formation of several subfragments. One of them exhibits the activity of FN-gelatinase that preferentially splits type I denatured collagen and fibronectin (see preceding paper). In this work we describe the purification and characterization of another fragment (25 kDa), issued from the same autodigest. This fragment may be activated to yield another proteinase, that splits preferentially laminin and denatured collagen type I. This enzyme will be referred as FN-laminase. Purified FN-laminase specifically reacted with antibodies against fibronectin. The specificity of bond cleavage by FN-laminase was studied with various synthetic peptides analogous to collagen repeats. FN-laminase cleaves the Ala-Gly bond in the sequence GPAGPR; the arginine residue in position P3&#39; is important for this cleavage. The enzyme is inhibited by pepstatin A and phenylmethanesulfonyl fluoride, like retroviral aspartic proteinases. It is also inhibited by EDTA. No inhibition was obtained with 1,10-phenanthroline or 4-chloromercuribenzoate, inhibitors of Zn-metalloproteinases or cysteine proteinases, respectively.

Latent fibronectin-degrading serine proteinase activity in N-terminal heparin-binding domain of human plasma fibronectin

European Journal of Biochemistry, 1991

The N-terminal 70-kDa fragment of human plasma fibronectin, purified from a cathepsin D digest, i... more The N-terminal 70-kDa fragment of human plasma fibronectin, purified from a cathepsin D digest, is characterized by lack of stability. It is processed proteolytically during incubation in the presence of Ca2+ into 27-kDa N-terminal heparin-binding and 45-kDa collagen-binding domains. The N-terminal residue in the 27-kDa fragment was blocked as in native fibronectin. The 45-kDa fragments began with the sequences AAVYQP, AVYQP and VYQP (residues 260, 261, 262-265 of fibronectin) that correspond to the beginning of the collagen-binding domain. In the presence of Ca2+ the purified 27-kDa fragment underwent further processing finally leading to the cleavage of the bond K85-D86 and to the simultaneous appearance of a specific proteolytic activity. Inhibition studies suggests that the newly generated enzyme is a Ca(2+)-dependent serine proteinase. Among all assayed matrix proteins, the newly generated enzyme cleaves native fibronectin and its fragments. It is proposed that this fibronectinase may originate from the N-terminal domain of fibronectin.

Potential proteolytic activity of human plasma fibronectin

Proceedings of the National Academy of Sciences, 1986

Evidence is presented that fibronectin (FN) polypeptide chain contains a latent proteinase. Human... more Evidence is presented that fibronectin (FN) polypeptide chain contains a latent proteinase. Human plasma FN was cleaved with cathepsin D into three main fragments: 140-kDa and 70-kDa single-chain and 140-kDa double-chain polypeptides. Their separation was achieved according to their affinity for heparin-Sepharose. A single-chain 140-kDa fragment (H-1) was eluted in the first peak. This peptide corresponds to the already described fragment that originates from the central part of FN; it contains a low-affinity heparinbinding site, one free SH group, and a cell-binding site. After reduction and further purification by preparative polyacrylamide gel electrophoresis, this fragment revealed a spontaneous decomposition, which could be attributed to proteolytic degradation. The subfragments, ranging from 25 to 95 kDa, yielded the same proteolytically active doublet of 28-30 kDa when tested by NaDodSO4/polyacrylamide gel electrophoresis in a gel containing copolymerized gelatin or fibrinogen. The proteolytic activity was inhibited by specific SH proteinase inhibitors. The proteinase forms a labeled complex after its incubation with '25I-labeled cystatin. Neither FN, cathepsin D, nor any products from previous purification steps were proteolytically active under the conditions of the assay. It was suggested that the same fragment may also yield an inhibitor, since structural analogies were found between the cell-binding region of FN and SH proteinase inhibitors.

Increasing cocoa butter yield through genetic improvement of Theobroma cacao L.: Seed fat content variability, inheritance, and association with seed yield

José Luis Pires1, Júlio Cézar Mattos Cascardo2, Smilja Vidmar Lambert2 & Antonio Figueira3 1 ... more

Activity evaluation of cocoa pod borer sex pheromone in cacao fields

Environmental …, 2008

Mention of trade names or commercial products in this article is solely for the purpose of provid... more

<I>Conopomorpha cramerella</I> (Lepidoptera: Gracillariidae) in the Malay Archipelago: Genetic Signature of a Bottlenecked Population?

Annals of the Entomological Society of America, 2008

Conopomorpha cramerella (Snellen) (Lepidoptera: Gracillariidae) is a devastating pest of cacao, T... more Conopomorpha cramerella (Snellen) (Lepidoptera: Gracillariidae) is a devastating pest of cacao, Theobroma cacao L. (Sterculiaceae), in Southeast Asia, particularly in the Malay Archipelago. We surveyed genetic variation at two unlinked loci, mitochondrial cytochrome oxidase I (COI) and nuclear elongation factor-1␣ (EF-1␣), in C. cramerella from throughout most of their known geographic range. Given the enormous area sampled, COI variation is extremely low; EF-1␣ variation may be low as well, but this is more difÞcult to assess due to the lack of appropriate data sets for comparison. Our results strongly suggest that sampled C. cramerella populations have experienced at least one bottleneck in their recent past, although the possibility that COI variation has been reduced by a selective sweep cannot be excluded based on available data. We suggest that one or more bottlenecks likely occurred when C. cramerella from an as yet unknown source population, either within or outside the Malay Archipelago, became established on cacao, which is not endemic to this region (Conopomorpha is an Old World genus and cacao originated in the New World). IdentiÞcation of the source of this pest could be important in efforts to identify natural enemies for biological control.

Testing local cocoa selections in three provinces in Sulawesi: (i) Productivity and resistance to cocoa pod borer and Phytophthora pod rot (black pod)

Crop Protection, 2015

ABSTRACT Trials were established on smallholder cocoa farms in three provinces in Sulawesi to ass... more ABSTRACT Trials were established on smallholder cocoa farms in three provinces in Sulawesi to assess productivity and constitutive responses of local cocoa clones to cocoa pod borer (CPB) and Phytophthora pod rot (Ppr) in different environmental situations. Twelve clones per trial (local farmer-assisted selections or clones produced by hybridisation programs in East Java and Malaysia) were tested in the districts of Pinrang, Polewali-Mandar and North Kolaka, including four standards common to the trials: the Malaysian clone, PBC123, and three selections from Sulawesi farms. The clones were evaluated from the time they started fruiting in 2010 (about two years after planting) for two years during which time chemical pesticides were not applied. Otherwise farms were managed according to recommended practices, including harvesting each fortnight, fertiliser application and heavy and light pruning, depending on the season. Butter fat content was generally lower than 50% but was higher in three local selections in Pinrang, M04, RB and Panimbu Red. While strongly dependent on genotype, fat and shell content and pod values in the common standards showed some variation between sites. The bean size and fat content of PBC123 was low, but this clone yielded better than most of the clones tested. For the common standards, yield estimates obtained from average yield per tree were higher in Pinrang (735-1100 tons/ha/annum) than in N. Kolaka (342-894 ton/ha/annum) or Polewali-Mandar (485-899 tons/ha/annum) indicating a marked site-effect. The number of flowers produced was higher in the common standards in Pinrang. Soil parameters including pH and exchangeable calcium, magnesium and potassium were higher in Pinrang than in Polman, although both sites were deficient in soil nitrogen and organic carbon. Lower average CPB infestation rates in ripe pods for the two-year evaluation period occurred in Pinrang (48-66%) and Polewali-Mandar (19-68%) than in N. Kolaka (77-80%). In most of the clones, total and severe CPB incidence decreased during the high pod season but some selections, such as M04 and TR01, maintained a low total and severe CPB incidence in both the low and high pod seasons, indicating partial resistance. In the ripe pods of common standards, the highest average Ppr incidence (ranging from 10 to 14%) occurred in N. Kolaka, which had a higher annual rainfall than the other sites. In ripe pods in the Pinrang trial, Geni J, M06 and Panimbu Red had a low Ppr incidence (4.4-4.8%) while M04 was Ppr-susceptible (23%). Incidence of Helopeltis spp. was high in the immature pods of some clones (exceeding 30% of the total harvest in M01 and Geni J in Pinrang). The results show that the performance of clones is affected by the locality in which they are grown, as well as their genotype, indicating the importance of testing clones under different environmental conditions. While the trials confirmed the efficacy of farmer-assisted selection, they also indicated that clones resistant to CPB, were susceptible to Ppr or other pests/diseases, and vice versa. For example, local selection, M04, was highly susceptible to Ppr, yet resistant to CPB. Therefore, the results indicate the importance of efforts to screen the progeny of hybrid crosses that combine resistance and yield traits.

Phosphonate applied by trunk injection controls stem canker and decreases< i> Phytophthora</i> pod rot (black pod) incidence in cocoa in Sulawesi

by David Guest and S. Lambert

Australasian Plant …, 2010

Stem canker and Phytophthora pod rot (PPR) or black pod caused by Phytophthora palmivora are seri... more Stem canker and Phytophthora pod rot (PPR) or black pod caused by Phytophthora palmivora are serious diseases of cocoa (Theobroma cacao L.) in Sulawesi, Indonesia, causing high yield losses for smallholders, possibly exceeded only by losses due to the cocoa pod borer (CPB), Conopomorpha cramerella. Potassium phosphonate (phosphite) applied by trunk injection has been demonstrated to effectively control canker and PPR in Papua New Guinea. The method was tested in a field trial in south-east Sulawesi. Fifty trees were injected with phosphonate, 50 with water and 50 were left untreated. Phosphonate was applied at a rate of~16 g active ingredient per tree per year, depending on the size of each tree. Trees were evaluated each month for canker severity, for PPR incidence and for CPB incidence and severity. From 4 months after the initial injection, trees treated with phosphonate had negligible levels of canker. Over a 2.5-year period, phosphonate significantly decreased PPR incidence. Cycles of PPR infection occurred in the wet season with PPR incidence fluctuating from less than 30% to greater than 75%. These fluctuations might have been due to variations in rainfall causing natural cycles of sporulation and infection. CPB incidence did not differ significantly between treatments. Since trunk injection of phosphonate effectively controls stem canker and decreases PPR in the long term it provides a valuable option for the management of these diseases for cocoa smallholders.

On-farm selection for quality and resistance to pest/diseases of cocoa in Sulawesi:(ii) quality and performance of selections against Phytophthora pod rot …

by David Guest, S. Lambert, Agung Susilo, and A. Iswanto

… Journal of Pest …, 2010

The cocoa industry in Sulawesi, the main region of cocoa production in Indonesia, is threatened b... more The cocoa industry in Sulawesi, the main region of cocoa production in Indonesia, is threatened by destructive diseases, including vascular-streak dieback (VSD) caused by the basidiomycete Oncobasidium theobromae and stem canker and Phytophthora pod rot (PPR) or black pod, caused by Phytophthora palmivora. Using the considerable genetic diversity of cocoa on farms, host resistance was identified and tested with the participation of farmers. Fortynine local and international cocoa selections with promising resistance characteristics (as well as susceptible controls) were side-grafted onto mature cocoa in a replicated trial with single-tree plots. Developing grafts were assessed in the dry season for severity of VSD infection, scored from 0 (no infection) to 4 (graft death). All of the 49 clones in the trial became infected with VSD in at least some replicates. Average severity varied from 0.2 to 1.6. Potential VSDresistance was found in eight clones, including DRC 15, KA2 106 and a local Sulawesi selection, VSD2Ldg. Some of the most susceptible clones were local Sulawesi selections from areas with a history of little or no VSD. Thirty-four pod-bearing clones were evaluated over a 2-year period for yield, quality and resistance to natural infections of PPR. Cumulative PPR incidence for all clones was 22% but varied from 8.6 to 43% among clones. Clones with less than 15% PPR incidence were designated as resistant, including DRC 16 and local Sulawesi selections, Aryadi 1, Aryadi 3 and VSD1Ldg. Scavina 12 was moderately resistant in the trial with a PPR incidence of 23%. Cumulative incidences of the mirid, Helopeltis spp., determined in the same evaluation period, indicated that DRC16 was the most susceptible clone with an incidence of 52% in ripe pods and 23% in immature pods. In comparison, KKM4 showed evidence of resistance to Helopeltis spp., with incidences of 34 and 0.8% in ripe and immature pods, respectively. The impact of diseases and pests (including cocoa pod borer) on bean losses and bean quality varied between clones but generally the bean size (or bean count) was affected more than the fat content or shell content.

On-farm selection for quality and resistance to pest/diseases of cocoa in Sulawesi: (i) performance of selections against cocoa pod borer, Conopomorpha cramerella

International Journal of Pest Management, 2009

2009)'On-farm selection for quality and resistance to pest/diseases of cocoa in Sulawesi: (i) per... more

Phosphonate applied by trunk injection controls stem canker and decreases Phytophthora pod rot (black pod) incidence in cocoa in Sulawesi

Australasian Plant Pathology, 2010

Stem canker and Phytophthora pod rot (PPR) or black pod caused by Phytophthora palmivora are seri... more Stem canker and Phytophthora pod rot (PPR) or black pod caused by Phytophthora palmivora are serious diseases of cocoa (Theobroma cacao L.) in Sulawesi, Indonesia, causing high yield losses for smallholders, possibly exceeded only by losses due to the cocoa pod borer (CPB), Conopomorpha cramerella. Potassium phosphonate (phosphite) applied by trunk injection has been demonstrated to effectively control canker and PPR in Papua New Guinea. The method was tested in a field trial in south-east Sulawesi. Fifty trees were injected with phosphonate, 50 with water and 50 were left untreated. Phosphonate was applied at a rate of~16 g active ingredient per tree per year, depending on the size of each tree. Trees were evaluated each month for canker severity, for PPR incidence and for CPB incidence and severity. From 4 months after the initial injection, trees treated with phosphonate had negligible levels of canker. Over a 2.5-year period, phosphonate significantly decreased PPR incidence. Cycles of PPR infection occurred in the wet season with PPR incidence fluctuating from less than 30% to greater than 75%. These fluctuations might have been due to variations in rainfall causing natural cycles of sporulation and infection. CPB incidence did not differ significantly between treatments. Since trunk injection of phosphonate effectively controls stem canker and decreases PPR in the long term it provides a valuable option for the management of these diseases for cocoa smallholders.

Vascular Streak Dieback of cacao in Southeast Asia and Melanesia: in planta detection of the pathogen and a new taxonomy

by Gary Samuels, Ade Rosmana, and S. Lambert

Fungal Biology, 2012

Keywords: Ceratobasidium Ceratobasidium ramicola Oncobasidium Plant pathology Thanatephorus Theob... more Keywords: Ceratobasidium Ceratobasidium ramicola Oncobasidium Plant pathology Thanatephorus Theobroma cacao Uthatobasidium a b s t r a c t Vascular Streak Dieback (VSD) disease of cacao (Theobroma cacao) in Southeast Asia and Melanesia is caused by a basidiomycete (Ceratobasidiales) fungus Oncobasidium theobromae (syn. ¼Thanatephorus theobromae). The most characteristic symptoms of the disease are green-spotted leaf chlorosis or, commonly since about 2004, necrotic blotches, followed by senescence of leaves beginning on the second or third flush behind the shoot apex, and blackening of infected xylem in the vascular traces at the leaf scars resulting from the abscission of infected leaves. Eventually the shoot apex is killed and infected branches die. In susceptible cacao the fungus may grow through the xylem down into the main stem and kill a mature cacao tree. Infections in the stem of young plants prior to the formation of the first 3e4 lateral branches usually kill the plant. Basidiospores released from corticioid basidiomata developed on leaf scars or along cracks in the main vein of infected leaves infect young leaves. The pathogen commonly infects cacao but there are rare reports from avocado. As both crops are introduced to the region, the pathogen is suspected to occur asymptomatically in native vegetation. The pathogen is readily isolated but cultures cannot be maintained. In this study, DNA was extracted from pure cultures of O. theobromae obtained from infected cacao plants sampled from Indonesia. The internal transcribed spacer region (ITS), consisting of ITS1, 5.8S ribosomal RNA and ITS2, and a portion of nuclear large subunit (LSU) were sequenced. Phylogenetic analysis of ITS sequences placed O. theobromae sister to Ceratobasidium anastomosis groups AG-A, AG-Bo, and AG-K with high posterior probability. Therefore the new combination Ceratobasidium theobromae is proposed. A PCR-based protocol was developed to detect and identify C. theobromae in Please cite this article in press as: Samuels GJ, et al., Vascular Streak Dieback of cacao in Southeast Asia and Melanesia: in planta detection of the pathogen and a new taxonomy, Fungal Biology (2011), doi:10.1016/j.funbio.2011.07.009 plant tissue of cacao enabling early detection of the pathogen in plants. A second species of

Cementing proteins provide extra mechanical stabilization to viral cages

The study of virus shell stability is key not only for gaining insights into viral biological cyc... more The study of virus shell stability is key not only for gaining insights into viral biological cycles but also for using viral capsids in materials science. The strength of viral particles depends profoundly on their structural changes occurring during maturation, whose final step often requires the specific binding of 'decoration' proteins (such as gpD in bacteriophage lambda) to the viral shell. Here we characterize the mechanical stability of gpD-free and gpD-decorated bacteriophage lambda capsids. The incorporation of gpD into the lambda shell imparts a major mechanical reinforcement that resists punctual deformations. We further interrogate lambda particle stability with molecular fatigue experiments that resemble the sub-lethal Brownian collisions of virus shells with macromolecules in crowded environments. Decorated particles are especially robust against collisions of a few k B T (where k B is the Boltzmann's constant and T is the temperature B300 K), which approximate those anticipated from molecular insults in the environment.

Potential proteolytic activity of human plasma fibronectin: fibronectin gelatinase

by xavier blondeau and S. Lambert

Biological Chemistry Hoppe-Seyler, 1990

Human plasma fibronectin contains a latent proteinase that after activation cleaves gelatin and f... more Human plasma fibronectin contains a latent proteinase that after activation cleaves gelatin and fibronectin. The autoactivation propensity of the two purified cathepsin D-produced fragments of fibronectin (190 and 120 kDa) was compared. Both polypeptides were spontaneously activated in the presence of Ca2+. This activation was inhibited by EDTA. The active gelatinase was isolated from the autodigest of the 190-kDa fragment. Among various protein substrates, including laminin and native type I and IV collagens, the purified enzyme degraded only gelatin and fibronectin. We have named this proteinase FN-gelatinase. FN-gelatinase is inhibited by phenylmethanesulfonyl fluoride and also by pepstatin A like retroviral aspartic proteinases. The amino-acid composition of the purified enzyme (35 kDa) was compared with the entire fibronectin sequence using the computer programme FIT. The optimal fit indicated that the 35-kDa fragment corresponds to the stretch # 1043-1404. This sequence contai...

Learning lessons from participatory breeding research with Sulawesi cocoa smallholders

by S. Lambert and Agung Susilo

The similarity of cocoa flavour of fermented seeds from fingerprinted genotypes of Theobroma cacao L. from Brazil and Malaysia

Fermentation of Theobroma cacao L. seeds has been considered to be the most important factor infl... more Fermentation of Theobroma cacao L. seeds has been considered to be the most important factor influencing cocoa flavour and has accordingly received most attention. Development of a test procedure in which small amounts of wet cocoa seeds in net bags are inserted in fermentations of wet seeds from single or mixed genotypes has allowed the flavour potential of a wide range of genotypes to be examined. Flavour profiles of cocoa liquors from genotypes grown at BAL Plantations Sdn Bhd, Malaysia differed consistently for various replicate preparations indicating a strong genetic contribution to flavour. In the present study, similar flavour profiles were developed from cocoa from the same genotypes grown in Brazil and Malaysia and subjected to the same conditions of post-harvest processing. Genetic fingerprinting by Rapid Amplified Polymorphic DNA (RAPD) analysis confirmed that most of the genotypes tested for flavour in Brazil and Malaysia were the same, although there were also a few er...

[Proteolytic potential of fibronectin and degradation of extracellular matrix]

Pathologie-biologie, 1990

Fibronectin is one of the major adhesive glycoproteins and bears interaction sites for both cell ... more Fibronectin is one of the major adhesive glycoproteins and bears interaction sites for both cell receptors and the extracellular matrix. Disappearance of fibronectin is the first step of cellular transformation in carcinogenesis. This phenomenon has been ascribed to increased proteolysis of fibronectin or of its cellular receptor. Results obtained during previous studies by the authors have shown that the fibronectin molecule has latent proteolytic activities which become apparent only after the action of other external proteases. Two proteinases, FN-gelatinase and FN-laminase, were identified in cathepsin D fibronectin digest. The acute activity of these two proteases is responsible for degradation of the extracellular matrix. Furthermore, the sequences and functions of both enzymes share a number of features with retroviral proteases.

project Improving cocoa production through farmer involvement in demonstration trials of potentially superior and pest/disease resistant genotypes and integrated management practices

by Jeff Neilon and S. Lambert

Potential Proteolytic Activity of Fibronectin: Fibronectin Laminase and its Substrate Specificity

Biological Chemistry Hoppe-Seyler, 1990

The purified 190-kDa fibronectin fragment produced by cathepsin D can be spontaneously activated ... more The purified 190-kDa fibronectin fragment produced by cathepsin D can be spontaneously activated in the presence of CaCl2. This activation generates new proteolytic activities and also results in the formation of several subfragments. One of them exhibits the activity of FN-gelatinase that preferentially splits type I denatured collagen and fibronectin (see preceding paper). In this work we describe the purification and characterization of another fragment (25 kDa), issued from the same autodigest. This fragment may be activated to yield another proteinase, that splits preferentially laminin and denatured collagen type I. This enzyme will be referred as FN-laminase. Purified FN-laminase specifically reacted with antibodies against fibronectin. The specificity of bond cleavage by FN-laminase was studied with various synthetic peptides analogous to collagen repeats. FN-laminase cleaves the Ala-Gly bond in the sequence GPAGPR; the arginine residue in position P3&#39; is important for this cleavage. The enzyme is inhibited by pepstatin A and phenylmethanesulfonyl fluoride, like retroviral aspartic proteinases. It is also inhibited by EDTA. No inhibition was obtained with 1,10-phenanthroline or 4-chloromercuribenzoate, inhibitors of Zn-metalloproteinases or cysteine proteinases, respectively.

Latent fibronectin-degrading serine proteinase activity in N-terminal heparin-binding domain of human plasma fibronectin

European Journal of Biochemistry, 1991

The N-terminal 70-kDa fragment of human plasma fibronectin, purified from a cathepsin D digest, i... more The N-terminal 70-kDa fragment of human plasma fibronectin, purified from a cathepsin D digest, is characterized by lack of stability. It is processed proteolytically during incubation in the presence of Ca2+ into 27-kDa N-terminal heparin-binding and 45-kDa collagen-binding domains. The N-terminal residue in the 27-kDa fragment was blocked as in native fibronectin. The 45-kDa fragments began with the sequences AAVYQP, AVYQP and VYQP (residues 260, 261, 262-265 of fibronectin) that correspond to the beginning of the collagen-binding domain. In the presence of Ca2+ the purified 27-kDa fragment underwent further processing finally leading to the cleavage of the bond K85-D86 and to the simultaneous appearance of a specific proteolytic activity. Inhibition studies suggests that the newly generated enzyme is a Ca(2+)-dependent serine proteinase. Among all assayed matrix proteins, the newly generated enzyme cleaves native fibronectin and its fragments. It is proposed that this fibronectinase may originate from the N-terminal domain of fibronectin.

Potential proteolytic activity of human plasma fibronectin

Proceedings of the National Academy of Sciences, 1986

Evidence is presented that fibronectin (FN) polypeptide chain contains a latent proteinase. Human... more Evidence is presented that fibronectin (FN) polypeptide chain contains a latent proteinase. Human plasma FN was cleaved with cathepsin D into three main fragments: 140-kDa and 70-kDa single-chain and 140-kDa double-chain polypeptides. Their separation was achieved according to their affinity for heparin-Sepharose. A single-chain 140-kDa fragment (H-1) was eluted in the first peak. This peptide corresponds to the already described fragment that originates from the central part of FN; it contains a low-affinity heparinbinding site, one free SH group, and a cell-binding site. After reduction and further purification by preparative polyacrylamide gel electrophoresis, this fragment revealed a spontaneous decomposition, which could be attributed to proteolytic degradation. The subfragments, ranging from 25 to 95 kDa, yielded the same proteolytically active doublet of 28-30 kDa when tested by NaDodSO4/polyacrylamide gel electrophoresis in a gel containing copolymerized gelatin or fibrinogen. The proteolytic activity was inhibited by specific SH proteinase inhibitors. The proteinase forms a labeled complex after its incubation with '25I-labeled cystatin. Neither FN, cathepsin D, nor any products from previous purification steps were proteolytically active under the conditions of the assay. It was suggested that the same fragment may also yield an inhibitor, since structural analogies were found between the cell-binding region of FN and SH proteinase inhibitors.

Increasing cocoa butter yield through genetic improvement of Theobroma cacao L.: Seed fat content variability, inheritance, and association with seed yield

José Luis Pires1, Júlio Cézar Mattos Cascardo2, Smilja Vidmar Lambert2 & Antonio Figueira3 1 ... more

Activity evaluation of cocoa pod borer sex pheromone in cacao fields

Environmental …, 2008

Mention of trade names or commercial products in this article is solely for the purpose of provid... more

<I>Conopomorpha cramerella</I> (Lepidoptera: Gracillariidae) in the Malay Archipelago: Genetic Signature of a Bottlenecked Population?

Annals of the Entomological Society of America, 2008

Conopomorpha cramerella (Snellen) (Lepidoptera: Gracillariidae) is a devastating pest of cacao, T... more Conopomorpha cramerella (Snellen) (Lepidoptera: Gracillariidae) is a devastating pest of cacao, Theobroma cacao L. (Sterculiaceae), in Southeast Asia, particularly in the Malay Archipelago. We surveyed genetic variation at two unlinked loci, mitochondrial cytochrome oxidase I (COI) and nuclear elongation factor-1␣ (EF-1␣), in C. cramerella from throughout most of their known geographic range. Given the enormous area sampled, COI variation is extremely low; EF-1␣ variation may be low as well, but this is more difÞcult to assess due to the lack of appropriate data sets for comparison. Our results strongly suggest that sampled C. cramerella populations have experienced at least one bottleneck in their recent past, although the possibility that COI variation has been reduced by a selective sweep cannot be excluded based on available data. We suggest that one or more bottlenecks likely occurred when C. cramerella from an as yet unknown source population, either within or outside the Malay Archipelago, became established on cacao, which is not endemic to this region (Conopomorpha is an Old World genus and cacao originated in the New World). IdentiÞcation of the source of this pest could be important in efforts to identify natural enemies for biological control.

Testing local cocoa selections in three provinces in Sulawesi: (i) Productivity and resistance to cocoa pod borer and Phytophthora pod rot (black pod)

Crop Protection, 2015

ABSTRACT Trials were established on smallholder cocoa farms in three provinces in Sulawesi to ass... more ABSTRACT Trials were established on smallholder cocoa farms in three provinces in Sulawesi to assess productivity and constitutive responses of local cocoa clones to cocoa pod borer (CPB) and Phytophthora pod rot (Ppr) in different environmental situations. Twelve clones per trial (local farmer-assisted selections or clones produced by hybridisation programs in East Java and Malaysia) were tested in the districts of Pinrang, Polewali-Mandar and North Kolaka, including four standards common to the trials: the Malaysian clone, PBC123, and three selections from Sulawesi farms. The clones were evaluated from the time they started fruiting in 2010 (about two years after planting) for two years during which time chemical pesticides were not applied. Otherwise farms were managed according to recommended practices, including harvesting each fortnight, fertiliser application and heavy and light pruning, depending on the season. Butter fat content was generally lower than 50% but was higher in three local selections in Pinrang, M04, RB and Panimbu Red. While strongly dependent on genotype, fat and shell content and pod values in the common standards showed some variation between sites. The bean size and fat content of PBC123 was low, but this clone yielded better than most of the clones tested. For the common standards, yield estimates obtained from average yield per tree were higher in Pinrang (735-1100 tons/ha/annum) than in N. Kolaka (342-894 ton/ha/annum) or Polewali-Mandar (485-899 tons/ha/annum) indicating a marked site-effect. The number of flowers produced was higher in the common standards in Pinrang. Soil parameters including pH and exchangeable calcium, magnesium and potassium were higher in Pinrang than in Polman, although both sites were deficient in soil nitrogen and organic carbon. Lower average CPB infestation rates in ripe pods for the two-year evaluation period occurred in Pinrang (48-66%) and Polewali-Mandar (19-68%) than in N. Kolaka (77-80%). In most of the clones, total and severe CPB incidence decreased during the high pod season but some selections, such as M04 and TR01, maintained a low total and severe CPB incidence in both the low and high pod seasons, indicating partial resistance. In the ripe pods of common standards, the highest average Ppr incidence (ranging from 10 to 14%) occurred in N. Kolaka, which had a higher annual rainfall than the other sites. In ripe pods in the Pinrang trial, Geni J, M06 and Panimbu Red had a low Ppr incidence (4.4-4.8%) while M04 was Ppr-susceptible (23%). Incidence of Helopeltis spp. was high in the immature pods of some clones (exceeding 30% of the total harvest in M01 and Geni J in Pinrang). The results show that the performance of clones is affected by the locality in which they are grown, as well as their genotype, indicating the importance of testing clones under different environmental conditions. While the trials confirmed the efficacy of farmer-assisted selection, they also indicated that clones resistant to CPB, were susceptible to Ppr or other pests/diseases, and vice versa. For example, local selection, M04, was highly susceptible to Ppr, yet resistant to CPB. Therefore, the results indicate the importance of efforts to screen the progeny of hybrid crosses that combine resistance and yield traits.

Phosphonate applied by trunk injection controls stem canker and decreases< i> Phytophthora</i> pod rot (black pod) incidence in cocoa in Sulawesi

by David Guest and S. Lambert

Australasian Plant …, 2010

Stem canker and Phytophthora pod rot (PPR) or black pod caused by Phytophthora palmivora are seri... more Stem canker and Phytophthora pod rot (PPR) or black pod caused by Phytophthora palmivora are serious diseases of cocoa (Theobroma cacao L.) in Sulawesi, Indonesia, causing high yield losses for smallholders, possibly exceeded only by losses due to the cocoa pod borer (CPB), Conopomorpha cramerella. Potassium phosphonate (phosphite) applied by trunk injection has been demonstrated to effectively control canker and PPR in Papua New Guinea. The method was tested in a field trial in south-east Sulawesi. Fifty trees were injected with phosphonate, 50 with water and 50 were left untreated. Phosphonate was applied at a rate of~16 g active ingredient per tree per year, depending on the size of each tree. Trees were evaluated each month for canker severity, for PPR incidence and for CPB incidence and severity. From 4 months after the initial injection, trees treated with phosphonate had negligible levels of canker. Over a 2.5-year period, phosphonate significantly decreased PPR incidence. Cycles of PPR infection occurred in the wet season with PPR incidence fluctuating from less than 30% to greater than 75%. These fluctuations might have been due to variations in rainfall causing natural cycles of sporulation and infection. CPB incidence did not differ significantly between treatments. Since trunk injection of phosphonate effectively controls stem canker and decreases PPR in the long term it provides a valuable option for the management of these diseases for cocoa smallholders.

On-farm selection for quality and resistance to pest/diseases of cocoa in Sulawesi:(ii) quality and performance of selections against Phytophthora pod rot …

by David Guest, S. Lambert, Agung Susilo, and A. Iswanto

… Journal of Pest …, 2010

The cocoa industry in Sulawesi, the main region of cocoa production in Indonesia, is threatened b... more The cocoa industry in Sulawesi, the main region of cocoa production in Indonesia, is threatened by destructive diseases, including vascular-streak dieback (VSD) caused by the basidiomycete Oncobasidium theobromae and stem canker and Phytophthora pod rot (PPR) or black pod, caused by Phytophthora palmivora. Using the considerable genetic diversity of cocoa on farms, host resistance was identified and tested with the participation of farmers. Fortynine local and international cocoa selections with promising resistance characteristics (as well as susceptible controls) were side-grafted onto mature cocoa in a replicated trial with single-tree plots. Developing grafts were assessed in the dry season for severity of VSD infection, scored from 0 (no infection) to 4 (graft death). All of the 49 clones in the trial became infected with VSD in at least some replicates. Average severity varied from 0.2 to 1.6. Potential VSDresistance was found in eight clones, including DRC 15, KA2 106 and a local Sulawesi selection, VSD2Ldg. Some of the most susceptible clones were local Sulawesi selections from areas with a history of little or no VSD. Thirty-four pod-bearing clones were evaluated over a 2-year period for yield, quality and resistance to natural infections of PPR. Cumulative PPR incidence for all clones was 22% but varied from 8.6 to 43% among clones. Clones with less than 15% PPR incidence were designated as resistant, including DRC 16 and local Sulawesi selections, Aryadi 1, Aryadi 3 and VSD1Ldg. Scavina 12 was moderately resistant in the trial with a PPR incidence of 23%. Cumulative incidences of the mirid, Helopeltis spp., determined in the same evaluation period, indicated that DRC16 was the most susceptible clone with an incidence of 52% in ripe pods and 23% in immature pods. In comparison, KKM4 showed evidence of resistance to Helopeltis spp., with incidences of 34 and 0.8% in ripe and immature pods, respectively. The impact of diseases and pests (including cocoa pod borer) on bean losses and bean quality varied between clones but generally the bean size (or bean count) was affected more than the fat content or shell content.

On-farm selection for quality and resistance to pest/diseases of cocoa in Sulawesi: (i) performance of selections against cocoa pod borer, Conopomorpha cramerella

International Journal of Pest Management, 2009

2009)'On-farm selection for quality and resistance to pest/diseases of cocoa in Sulawesi: (i) per... more

Phosphonate applied by trunk injection controls stem canker and decreases Phytophthora pod rot (black pod) incidence in cocoa in Sulawesi

Australasian Plant Pathology, 2010

Stem canker and Phytophthora pod rot (PPR) or black pod caused by Phytophthora palmivora are seri... more Stem canker and Phytophthora pod rot (PPR) or black pod caused by Phytophthora palmivora are serious diseases of cocoa (Theobroma cacao L.) in Sulawesi, Indonesia, causing high yield losses for smallholders, possibly exceeded only by losses due to the cocoa pod borer (CPB), Conopomorpha cramerella. Potassium phosphonate (phosphite) applied by trunk injection has been demonstrated to effectively control canker and PPR in Papua New Guinea. The method was tested in a field trial in south-east Sulawesi. Fifty trees were injected with phosphonate, 50 with water and 50 were left untreated. Phosphonate was applied at a rate of~16 g active ingredient per tree per year, depending on the size of each tree. Trees were evaluated each month for canker severity, for PPR incidence and for CPB incidence and severity. From 4 months after the initial injection, trees treated with phosphonate had negligible levels of canker. Over a 2.5-year period, phosphonate significantly decreased PPR incidence. Cycles of PPR infection occurred in the wet season with PPR incidence fluctuating from less than 30% to greater than 75%. These fluctuations might have been due to variations in rainfall causing natural cycles of sporulation and infection. CPB incidence did not differ significantly between treatments. Since trunk injection of phosphonate effectively controls stem canker and decreases PPR in the long term it provides a valuable option for the management of these diseases for cocoa smallholders.

Vascular Streak Dieback of cacao in Southeast Asia and Melanesia: in planta detection of the pathogen and a new taxonomy

by Gary Samuels, Ade Rosmana, and S. Lambert

Fungal Biology, 2012

Keywords: Ceratobasidium Ceratobasidium ramicola Oncobasidium Plant pathology Thanatephorus Theob... more Keywords: Ceratobasidium Ceratobasidium ramicola Oncobasidium Plant pathology Thanatephorus Theobroma cacao Uthatobasidium a b s t r a c t Vascular Streak Dieback (VSD) disease of cacao (Theobroma cacao) in Southeast Asia and Melanesia is caused by a basidiomycete (Ceratobasidiales) fungus Oncobasidium theobromae (syn. ¼Thanatephorus theobromae). The most characteristic symptoms of the disease are green-spotted leaf chlorosis or, commonly since about 2004, necrotic blotches, followed by senescence of leaves beginning on the second or third flush behind the shoot apex, and blackening of infected xylem in the vascular traces at the leaf scars resulting from the abscission of infected leaves. Eventually the shoot apex is killed and infected branches die. In susceptible cacao the fungus may grow through the xylem down into the main stem and kill a mature cacao tree. Infections in the stem of young plants prior to the formation of the first 3e4 lateral branches usually kill the plant. Basidiospores released from corticioid basidiomata developed on leaf scars or along cracks in the main vein of infected leaves infect young leaves. The pathogen commonly infects cacao but there are rare reports from avocado. As both crops are introduced to the region, the pathogen is suspected to occur asymptomatically in native vegetation. The pathogen is readily isolated but cultures cannot be maintained. In this study, DNA was extracted from pure cultures of O. theobromae obtained from infected cacao plants sampled from Indonesia. The internal transcribed spacer region (ITS), consisting of ITS1, 5.8S ribosomal RNA and ITS2, and a portion of nuclear large subunit (LSU) were sequenced. Phylogenetic analysis of ITS sequences placed O. theobromae sister to Ceratobasidium anastomosis groups AG-A, AG-Bo, and AG-K with high posterior probability. Therefore the new combination Ceratobasidium theobromae is proposed. A PCR-based protocol was developed to detect and identify C. theobromae in Please cite this article in press as: Samuels GJ, et al., Vascular Streak Dieback of cacao in Southeast Asia and Melanesia: in planta detection of the pathogen and a new taxonomy, Fungal Biology (2011), doi:10.1016/j.funbio.2011.07.009 plant tissue of cacao enabling early detection of the pathogen in plants. A second species of