Cadmium (Cd), a toxic heavy metal found in agricultural landscapes worldwide, has been pointed out in cropped soils with Theobroma cacao L., as one of the main contaminants that translocate into plant tissues. Among the factors linked to cadmium translocation into plants, the role of soil bacterial communities in chemical transformation in soils has been poorly investigated. Overall, soil bacterial communities are shaped by diverse environmental and anthropogenic factors that influence crop yield and health. Cadmium alters soil microbial communities and increases the risk to human health through plant uptake. Although the impacts of cadmium on soil bacteria have been studied in other crops, there is limited information on cacao. Thus, this study aimed to assess the responses of soil bacterial communities in cacao farms to cadmium exposure, both natural and spiked. A total of 225 rhizosphere soil samples were collected from 16 plots across five cacao farms in two Colombian departments. The complementary approaches used were: (i) 16S rDNA amplicon sequencing to assess the composition of the bacterial community in soils with natural Cd concentrations, and (ii) isothermal microcalorimetry (IMC) to measure the temporal metabolic responses of bacteria to Cd in closed systems for 80 hours at 25 °C. The findings suggest that nearly 28% of the bacterial community responds to high cadmium concentrations in soils, both in natural and experimental conditions. Field-based observations revealed that Cd-responsive taxa detected under natural soil conditions included several unculturable bacterial groups, whereas laboratory experiments with Cd spiking predominantly selected for previously characterized cadmium-tolerant bacteria (CdtB). Significant variation in natural Cd-bacterial community composition and Cd-related metabolic activity was observed across the farms. Moreover, Cd-responsive bacterial taxa exhibited increased abundance during Cd spikes. As expected, contrasting patterns were revealed by the activity-response measured by IMC and taxonomic analyses of 16S rRNA gene sequences.

A mitigation strategy involving the application of cadmium-tolerant bacteria (CdtB) was assessed in cacao soils in farms in Arauca, Colombia. The bacterial strain used, Pseudomonas chlororaphis CdtBSO has exhibited an ability to immobilise cadmium, reducing its bioavailability to cacao. The aim of this study was to increase the immobilization activity of CdtB in cacao soils in Arauca. Five treatments combining different concentrations of CdtB with a synthetic fertiliser (Agrocacao) and zeolite were established. The cadA gene as marker for Cd immobilization was detected using droplet digital PCR (ddPCR). Moreover, the metabolic activity of CdtB was assessed using isothermal microcalorimetry (IMC). Maximum thermal performance was observed in the treatments where the most CdtB was applied. However, the results were highly dependent on the site. An increase in soil Cd content six months after application of the treatments was observed suggesting increased immobilization rates of the metal in the soil. The explanation of possible immobilisation mechanisms varied between treatments. However, the treatment showing the most positive trends included 50% CdtB and 50% Agrocacao. The detection of the cadA gene, revealed variations in the relative abundance of the copies of the gene in the assessed soils. This work contributes to the knowledge on soil Cd bioremediation using CdtB, through the application of innovative approaches to monitoring, such as ddPCR and IMC. It enriches the information available on the application of physicochemical and biological amendments for the remediation of Cd in real cacao crop conditions.

Cadmium contamination in cacao crops from Arauca, Colombia, is a major concern for exports. One of the causes is the contaminated fertilisers or the lack of a fertilisation scheme. This study assesses a pilot-field application of a chemical fertiliser (Agrocacao), zeolite, and a cadmium-tolerant bacterial (CdtB) formulation developed by AGROSAVIA. The treatments comprised combinations and varying proportions of the three amendments, a full-dose treatment, and an untreated control. The treatments were administered over 2 years, with a dosage applied twice a year. The Cd content was tested in soil and cacao leaves. The best treatments were identified for each location; however, no single treatment proved to be the best across all locations. Nonetheless, treatments T5 and T8 showed a significant positive trend for soil Cd (p = 0.124) and leaf Cd (p = 0.056) at two of the four locations tested in soil and leaves, respectively. These treatments included Agrocacao and Cd-tolerant bacteria (CdtB). The soil Cd concentrations ranged from 0.1 to 5.6 mg kg−1, whereas the leaf Cd concentrations varied from 0.2 to 38 mg kg−1. Cd immobilisation in soils reached up to 80

The cacao crop is important for Colombia due to its high economic impact. One concern is the presence of cadmium (Cd) in beans, which can enter the soil from various sources, including fertilisers. However, to date, no studies have been carried out to identify Cd in these inputs. Therefore, this study seeks to determine Cd concentrations in (i) organic matter, (ii) chemical fertilisers and (iii) lime, together with the available fraction accessible to the cacao tree. A total of 133 fertilisers used in eight departments of Colombia were assessed. For 10 of these fertilisers a sequential extraction of Cd was carried out in five categories: water-soluble, exchangeable, reducible, oxidizable and residual fractions. Likewise, the mobility of available Cd from the fertiliser to the soil was assessed with an incubation experiment. This study showed that organic fertilisers had the highest total Cd content of all those studied, and increased the Cd retained in the soils. In contrast, the mobile fraction and bioavailable Cd were higher in chemical fertilisers. These findings show a need to raise awareness about the rational use of fertilisers, as well as greater monitoring by local government authorities and elaboration of new regulations for maximum limits.

Assessing the bacterial community composition across cacao crops is important to understand its potential role as a modulator of cadmium (Cd) translocation to plant tissues under field conditions; Cd mobility between soil and plants is a complex and multifactorial problem that cannot be captured only by experimentation. Although microbes have been shown to metabolize and drive the speciation of Cd under controlled conditions, regardless of the link between soil bacterial community (SBC) dynamics and Cd mobilization in the rhizosphere, only a few studies have addressed the relationship between soil bacterial community composition (SBCC) and Cd content in cacao seeds (Cdseed). Therefore, this study aimed to explore the association between SBCC and different factors influencing the distribution of Cd across cacao crop systems. This study comprised 225 samples collected across five farms, where we used an amplicon sequencing approach to characterize the bacterial community composition. The soil Cd concentration alone (Cdsoil) was a poor predictor of Cdseed. Still, we found that this relationship was more apparent when the variation within farms was controlled, suggesting a role of heterogeneity within farms in modulating Cd translocation and, thus, seed Cd content. Our results provide evidence of the link between soil bacterial communities and the distribution of Cd across Colombian cacao crops, and highlight the importance of incorporating fine-spatial-scale studies to advance the understanding of factors driving Cd uptake and accumulation in cacao plants. IMPORTANCE:Cadmium (Cd) content in cacao crops is an issue that generates interest due to the commercialization of chocolate for human consumption. Several studies provided evidence about the non-biological factors involved in its translocation into the cacao plant. However, factors related to this process, including soil bacterial community composition (SBCC), still need to be addressed. It is well known that soil microbiome could impact compounds' chemical transformation, including Cd, on the field. Here, we found the first evidence of the link between soil bacterial community composition and Cd concentration in cacao soils and seeds. It highlights the importance of including the variation of bacterial communities to assess the factors driving the Cd translocation into cacao seeds. Moreover, the results highlight the relevance of the spatial heterogeneity within and across cacao farms, influencing the variability of Cd concentrations.

Cacao crops in Colombia play a fundamental role in its economy. Cacao bean exports have a rising tendency due to increased area and production. However, cadmium (Cd) in cacao beans has affected market in Colombia. The aim of this study is to assess Cd distribution hotspots for cacao beans, Cd in beans and soil relationships, geological ages and soil chemical parameters, overall Cd dynamics, soil-to-plant translocation, and EU regulation impacts on Cd presence in Colombian cacao production. A hot and cold spot analysis combined with a cluster and outlier analysis was used to build the geographic Cd distribution in cacao beans in Colombia. Cadmium in the subsurface and the relationship with Cd in the soil surface of cacao farms were analysed using the geoaccumulation index (Igeo). The bioconcentration factor (BCF) and soil chemical features in cacao beans were quantified. Positive correlations were observed within all the studied variables except for Mn and Fe. The highest correlation was observed within Ca, pH, and Cd in soil. The Valanginian-Albian and Albian-Maastrichtian geological ages are directly related to Cd in beans, Zn, P, soil Cd, and Fe. Regions with the highest cacao productivity and export registered an average Cd concentration in beans of 2.39_ ± _2.82 mg kg-1. Two municipalities in Santander were 'partially contaminated' according to Igeo. The BCF was not directly proportional to soil and bean Cd level and did not fully coincide with hot spots. This evidence reinforces the idea that even areas with apparently low Cd levels may register a latent problem associated with genetic/ontogenetic, climatic, or management factors. This first distribution map of Cd in cacao beans constitutes a tool to implement actions to minimize risks and face challenges posed by the imposed regulations that may limit raw cacao bean exports in Colombia.

The district of Arauca is the second-largest producer of cacao in Colombia. However, despite its quality, it faces issues for export due to levels of cadmium (Cd) higher than the regulatory thresholds. A central question is how it may impact agricultural performance in the presence of Cd in cacao and chocolates. This study quantified Cd in cacao plantations from Arauca. Thus, 180 farms were assessed in the municipalities of Arauquita, Fortul, Saravena, and Tame. Five sample types (soil, irrigation channel sediment, soil litter, cacao seeds, and chocolates) were assessed for Cd. As a technological innovation, the new MXRF technology was used for Cd in chocolates. The sequence of Cd content was soil litter > chocolate > soils > cacao seeds > irrigation-channel sediment. A gradient north–south of Cd content in soil was observed, where highest content was found in farms near the Arauca River, and lower farther away. In irrigation channel sediment, Cd levels averaged 0.07 mg kg −1 . The Cd content in cacao seeds was 0.78 mg kg −1 on average. Cd content in chocolates was above the threshold (1.10 mg kg −1 on average, including several cacao mass percentages). These artisanal chocolate bars produced by single farms were near the limit of Cd set by the European Union (up to 0.8 mg kg −1 ). Therefore, mixing beans from different farms could reduce their Cd content. The present study underscores the complexity of Cd distribution, emphasizing the importance of integrating soil, crop, and landscape features in managing and mitigating Cd levels in cacao.

The presence of Staphylococcus aureus in six dry-cured meat-processing facilities was investigated. S. aureus was detected in 3.8% of surfaces from five facilities. The occurrence was clearly higher during processing (4.8%) than after cleaning and disinfection (1.4%). Thirty-eight isolates were typified by PFGE and MLST. Eleven sequence types (STs) were defined by MLST. ST30 (32%) and ST12 (24%) were the most abundant. Enterotoxin genes were detected in 53% of isolates. The enterotoxin A gene (sea) was present in all ST30 isolates, seb in one ST1 isolate, and sec in two ST45 isolates. Sixteen isolates harbored the enterotoxin gene cluster (egc) with four variations in the sequence. The toxic shock syndrome toxin gene (tst) was detected in 82% of isolates. Regarding antimicrobial resistance, twelve strains were susceptible to all the antibiotics tested (31.6%). However, 15.8% were resistant to three or more antimicrobials and, therefore, multidrug-resistant. Our results showed that in general, efficient cleaning and disinfection procedures were applied. Nonetheless, the presence of S. aureus with virulence determinants and resistance to antimicrobials, particularly multidrug-resistant MRSA ST398 strains, might represent a potential health hazard for consumers.

Cacao (Theobroma cacao L) is one of the most relevant crops in terms of economy and social rural development in Colombia. Cacao is also an important crop due to its potential to replace illicit crops and it is related to less deforestation and preserves the biodiversity. There are several cacao districts in Colombia, one of these being Arauca. The Department of Arauca is the second largest cocoa producing region in Colombia; however, it is heavily affected by armed conflict. To raise the knowledge and technology available in the region, integrating data on the occurrence of cacao farms with climatic variables becomes a powerful socioeconomic mapping tool for maintaining agrobiodiversity and food security in the region. Consequently, this type of agrodiversity data and agroclimatic approaches help to better manage agrobiodiversity, as in the cacao region of Arauca. These tools are even more relevant in biodiverse regions, such as flooded savannahs and tropical forest ecosystems, which are currently undergoing drastic changes due to agricultural expansion and climate change. One of the knowledge gaps in Colombia´s cacao regions is that there are currently no agroclimatic maps made with a social and scientific approach. This study aimed to provide a database of the spatial distribution of cacao farms in Arauca, as well as agroclimatic maps that identify and locate cacao climate regions in Arauca. We also present a presence-only matrix consisting of twenty-six tree species, or agrobiodiversity, distributed across the study region and specifically associated with the cacao forestry systems in Arauca.We present the first database of both climate and agrobiodiversity data related to cacao farms in Arauca, developed with a research and socioeconomic vision that generated a novel approach for the agroclimatic zoning of cocoa in the Arauca Region and Colombia. Using 1,538 cacao farms at the regional scale, we identified two national and six regional-scale climate and soil regions. The selection at the local scale allowed us to classify 180 cacao farms comprising nine agroclimatic clusters in Arauca. We found twenty-six tree species distributed across the cacao climate zones. This dataset and its related maps also represent the agrobiodiversity of cultivated cacao locally. This is the most complete climate and agrobiodiversity dataset of cacao farms distribution in one of the top cocoa-producing regions in the country. These outputs are crucial because they constitute a baseline for developing research in the biodiversity of agroforestry systems, pests and diseases, pollutant presence, genetics, post-harvest processing and cocoa quality and safety.

In cacao farms, the presence of cadmium (Cd) is a major issue for commercialization, particularly for countries such as Colombia. Cadmium-tolerant bacteria (CdtB) are an important functional group of microorganisms with a potential for bioremediation strategies. Cd immobilization activity by CdtB can be accurately measured by isothermal microcalorimetry (IMC). In this study, the metabolic capacity of an entire CdtB community in cacao farm soils from three cacao-producing districts of Colombia, with and without the addition of Cd was measured using IMC. The differences between the observed peaks in metabolic activity related to Cd immobilization were analysed to determine whether activation of CdtB populations occurred when Cd content was increased. The thermograms from Santander soil samples have a major metabolic activity of the CdtB community compared to peaks of maximal heat-flow in the samples from Antioquia and Arauca. IMC showed differences in Cd immobilization ratios of the soil samples of 0.11–0.30 mg kg−1 h−1 at 25°C over 12 days of thermal monitoring. Furthermore, the amplicons of cadA and smt genes from the soil samples allow elucidation of possible metabolic mechanisms used by CdtB soil populations. The gene amplification confirmed the existence of CdtB populations related to both bioweathering and biochelating metabolic capacities. Scanning electron microscope (SEM) images supported the existence of otavite biologically induced by CdtB naturally. A Pearson correlation analysis was made between kinetical growth parameters and thermodynamic data. Besides, a PCA was performed between CdtB cadA gene copies, soil pH and SOM indicating the effect of CdtB in Cd translocation. Thus, it is concluded that the combination of Cd immobilization ratios obtained using isothermal microcalorimetry, the molecular basis of metabolic immobilization, and SEM imagery could act as a useful toolkit to identify CdtB populations for bioremediation strategies in contaminated cacao farms. The research importance of this study is the use of combined tools for quantitative IMC measurements to identify and assess Cd metabolic capacities of CdtB populations in soil, in situ, as a new proxy for CdtB assessment in cacao-growing soils.

In-situ geophysical techniques based on electrical resistivity tomography were performed to determine the spatial and temporal distribution of heavy metals. This paper aimed to pursue a novel methodology consisting in combining electrical resistivity tomography measurements, soil chemical data, and statistical analysis to predict As and Cd distribution in paddy soils. The resistivity surveys were performed at 2 m depth in nine locations of Tolima district in Colombia. Chemical concentrations of both As and Cd were introduced in a standardization procedure to estimate their electrical resistivity values. Soil and fertilizers information were acquired to determine As and Cd input in the studied area. The laboratory standardization showed a direct linear regression for Cd and an inverse one for As. A high positive Spearman correlation between electrical resistivity and As / Cd content was determined, with 0.98 and 0.99 values, respectively. Results showed that As-like compounds were distributed mainly in shallower soil layers (0.45 m) and were characterized by lower electrical resistivity values (181-2318 omega.m). On the contrary, Cd-like compounds were observed in a deeper profile (0.80 m) with higher electrical resistivity values (559-4879 omega.m). This work underlies the utility of the combined geophysical techniques, soil chemical parameters and statistical analysis to improve the understanding of As and Cd dynamics and origin in topsoil, supporting the need of an agricultural management of paddy soils.

Bioremediation of farm soil is a technique that merits in-depth research. There are few studies related to the use of bioremediation to reduce cadmium (Cd) availability in soils used for cacao production. This study investigates (1) field bioprospection and strain characterization using techniques including isothermal microcalorimetry to select a group of cadmium-tolerant bacteria (CdtB) for potential use as bioremediators of cacao soils and (2) the application of bacterial inoculum to compare the immobilization of Cd under field conditions. Bioprospection was carried out in four cacao farms from the Antioquia district in Colombia. Culturable CdtB strains were isolated using CdCl2 as a Cd source and identified using molecular techniques. The metabolic characterization of Cd immobilization was carried out using isothermal microcalorimetry with CdCl2 amendments. Five cadmium-tolerant bacteria were isolated and characterized as Bacillus spp. The strain CdtB14 showed better growth and Cd immobilization ability (estimated through heat ratios) than any strain isolated thus far, suggesting potential for future use in bioproduct development. Furthermore, the application of two previously characterized CdtB strains with zeolite powder was performed in the same farms where the bioprospection process was carried out. The application of the preformulated inoculum resulted in a decrease of 0.30 + 0.1 mg kg−1 of soil Cd in two out of the four assessed farms. The field results are preliminary and require data on the change in Cd in cacao beans to understand what this result means for Cd mitigation. This study is the first to combine bioprospecting and the performance of CdtB in laboratory and field experiments in cacao farms and shows the potential of bioremediation to mitigate Cd contamination in cacao.

Abstract Regulation of maximum levels of cadmium in chocolate is an issue for cacao exportation from many parts of Latin America, including Colombia. These limits are related to the final product, but buyers often request maximum levels of Cd in the beans. However, to date, there is neither a clear understanding of the relationship between the specified levels of Cd in chocolate and cocoa derivatives and levels in harvested beans or soil nor of the effect of post-harvest processes on the levels of Cd in the final product. To address this, the fate of Cd concentration from soil to chocolate bar was followed in a single farm in Santander district, Colombia. The concentration of Cd in soils was measured using ICP-OES and correlated with soil pH, soil organic matter (SOM), and the use of P-based fertilisers. Cd concentrations were also measured in unfermented seeds, fermented and dried beans, shell, nibs, and chocolate. SOM (2.93–3.78%), soil pH (4.7–4.9), soil P concentration (120–132 mg kg−1) affect Cd availability. However, it is still unclear whether Cd concentration of P-based fertilisers (3–30 mg kg−1) is important or not. While post-harvest treatments did not affect the Cd concentration of beans (4.17 ± 0.8 mg kg−1 on average), the removal of the shell (6.57 mg kg−1) from the nibs (3.28 mg kg−1), as well as the percentage of cocoa mass used contributes to a reduction in Cd concentration in the chocolate bar (1.60 mg kg−1). This study provides clear indications on where research into mitigation measures should be focussed, as well as indicating the importance of carrying out analyses for Cd in the nib or cacao mass, rather than the whole bean, reducing Cd concentration by up to 40%. Graphical Abstract

Cadmium (Cd) is considered a toxic heavy metal; nevertheless, its toxicity fluctuates for different organisms. Cadmium-tolerant bacteria (CdtB) are diverse and non-phylogenetically related. Because of their ecological importance these bacteria become particularly relevant when pollution occurs and where human health is impacted. The aim of this review is to show the significance, culturable diversity, metabolic detoxification mechanisms of CdtB and their current uses in several bioremediation processes applied to agricultural soils. Further discussion addressed the technological devices and the possible advantages of genetically modified CdtB for diagnostic purposes in the future.

Cadmium (Cd) is a non-nutritive element present across the earth. In cacao crops from South America, Cd has become one of the biggest challenges due to its flux from soils, and due to the enriched content of this metal, it makes its way into the beans and finally affects the chocolate quality. This manuscript aims to show how the flux of Cd occurs, from the soil to the post-harvest phase and chocolate production, by analysing the possible inputs to the system in a single farm assessed as a model for enriched-Cd status. This study shows that both geogenic and anthropogenic activities have an incidence on the final Cd content in chocolate, especially with respect to soil properties, fertiliser applications, post-harvest treatments and chocolate production.

Esta serie de cartillas son importantes para el cacaocultor porque muestran varios aspectos que se deben tener en cuenta en el cultivo de cacao para la prevención y reducción de cadmio. La primera cartilla aborda los conceptos básicos, definiendo el cadmio y explicando su presencia en la naturaleza y su origen. También se describe cómo este elemento afecta la salud humana, cuál es la normativa internacional sobre el cadmio en el cacao y los criterios de manejo de cadmio en la industria cacaotera. La segunda cartilla presenta los aportes de investigación y desarrollo de varias instituciones en Colombia por mitigar o disminuir la presencia de cadmio en el cacao y aborda temas como fertilización, enmiendas y microorganismos. Por último, la tercera cartilla da cuenta del plan de acción del Gobierno nacional y de todas las instituciones que de alguna manera están trabajando de forma unida y alineada con otros actores de la cadena para ir mejorando la problemática de la presencia de cadmio en algunos sitios de Colombia.

This study represents the first nationwide survey regarding the distribution of Cd content in cacao-growing soils in Colombia. The soil Cd distribution was analyzed using a cold/hotspots model. Moreover, both descriptive and predictive analytical tools were used to assess the key factors regulating the Cd concentration, considering Cd content and eight soil variables in the cacao systems. A critical discussion was performed in four main cacao-growing districts. Our results suggest that the performance of a model using all the variables will always be superior to the one using Zn alone. The analyzed variables featured an appropriate predictive performance, nonetheless, that performance has to be improved to develop a prediction method that might be used nationwide. Results from the fitted graphical models showed that the largest associations (as measured by the partial correlation coefficients) were those between Cd and Zn. Ca had the second-largest partial correlation with Cd and its predictive performance ranked second. Interestingly, it was found that there was a high variability in the factors correlated with Cd in cacao growing soils at a national level. Therefore, this study constitutes a baseline for the forthcoming studies in the country and should be reinforced with an analysis of cadmium content in cacao beans.

Cadmium is a toxic non-essential metal for almost all living systems. It is one of the biggest challenges for farmlands and the food chain because of its toxic effect on human health. This research aims to determine the Cd content in soils, litter and cacao beans, following the Cd fluxes within each cacao system using the two-dimensional electrical resistivity tomography technique. The study was carried out in four farms located in the Magdalena basin in Antioquia, Colombia. The farms showed a heterogeneity in relation to cacao cultivars, altitude, topography and geology. The soil cation electrical capacity, pH and soil organic matter levels, as well as the Al, Al3+H+, Ca, K, Mg and P contents were measured at different depths. Moreover, the Cd content was correlated to the resistivity values of samples taken in situ using ERT. Soil Al3+H+ and the altitude of the farms fitted as the best predictors of the beans’ Cd content. Furthermore, the Cd content in soils from the assessed farms ranged between 1.22 and 2.03 mg kg−1. The Cd content in cocoa beans ranged from 0.07 to 1.44 mg kg−1, with a value of 0.40 mg kg−1 on average. The resistivity values obtained in field showed a high correlation with the soil Cd content determined (R2 = 0.82). The predictive tomography plots highlighted topsoil Cd dynamics between litter, amendments and fertilizers. Therefore, these results underlie the utility of the combined geophysical techniques and soil chemical properties including the analysis of fertilizer amendments to improve the understanding of Cd dynamic.

Esta serie de cartillas son importantes para el cacaocultor porque muestran varios aspectos que se deben tener en cuenta en el cultivo de cacao para la prevención y reducción de cadmio. La primera cartilla aborda los conceptos básicos, definiendo el cadmio y explicando su presencia en la naturaleza y su origen. También se describe cómo este elemento afecta la salud humana, cuál es la normativa internacional sobre el cadmio en el cacao y los criterios de manejo de cadmio en la industria cacaotera. La segunda cartilla presenta los aportes de investigación y desarrollo de varias instituciones en Colombia por mitigar o disminuir la presencia de cadmio en el cacao y aborda temas como fertilización, enmiendas y microorganismos. Por último, la tercera cartilla da cuenta del plan de acción del Gobierno nacional y de todas las instituciones que de alguna manera están trabajando de forma unida y alineada con otros actores de la cadena para ir mejorando la problemática de la presencia de cadmio en algunos sitios de Colombia.