Determination of ochratoxin A in cocoa beans using immunoaffinity column cleanup with high-performance liquid chromatography.

A method was developed and validated for the determination of ochratoxin A (OTA), a fungal metabolite, in cocoa beans of high fat content. The sample was extracted by blending with a 1% sodium bicarbonate solution (pH 10) followed by ultrasonication, and the sample was defatted by treatment with a flocculant. The defatted sample was purified using immunoaffinity column chromatography, and OTA was detected using HPLC with fluorescence detection. The method was fully optimized, validated, and quality controlled using spike recovery analyses, with recoveries of 89-105% over spiking ranges of 320-2.5 ng/g with CV of analyses generally <10% over 4 consecutive years and an LOQ of 0.66 ng/g in cocoa bean samples. This method overcomes the problems posed by the high fat contents of cocoa and chocolate samples with a high degree of reliability.

Modelling response patterns of physico-chemical indicators during high-rate composting of green waste for suppression of Pythium ultimum.

High-rate composting studies on green waste, i.e. banana leaves (BL) and lawn clippings (LC), were conducted in 0.25-m3 rotary barrel composters to evaluate and model changes in key physico-chemical parameters during composting. Time to compost maturity and antagonistic effects and relationships of composts against Pythium ultimum were also investigated. Higher temperatures were achieved in LC compost (LCC), which did not translate to higher total organic carbon (TOC) loss but resulted in lower carbon to nitrogen ratio (C:N) and a more mature compost. With the exception of electrical conductivity (EC), net decreases were observed in pH, TOC and C:N across compost types. Total Kjeldahl nitrogen (TKN) showed a net increase in LCC and a net decrease in BLC. With the exception of TOC and pH, the results showed that compost type and time had a significant effect on the respective TKN, EC and C:N models. Compost temperature and TOC were best described by the critical exponential and rectangular hyperbola functions, respectively. Whereas TKN, C:N and pH were described using double Fourier functions and EC using Fourier functions. Composts achieved maturity within 19 days and significantly inhibited the growth of P. ultimum. Bacterial population was positively related to growth inhibition (GI) across compost types, whereas total microbial population had a positive relationship with GI in LCC. Evidence suggests that multiple groups of microorganisms contributed to GI through antibiosis and competition for resources. Composts were determined to be suitable for use as components of plant growth substrates based on compost maturity indices.

Genome-wide association studies and genomic selection assays made in a large sample of cacao (Theobroma cacao L.) germplasm reveal significant marker-trait associations and good predictive value for improving yield potential.

A genome-wide association study (GWAS) was undertaken to unravel marker-trait associations (MTAs) between SNP markers and phenotypic traits. It involved a subset of 421 cacao accessions from the large and diverse collection conserved ex situ at the International Cocoa Genebank Trinidad. A Mixed Linear Model (MLM) in TASSEL was used for the GWAS and followed by confirmatory analyses using GAPIT FarmCPU. An average linkage disequilibrium (r2) of 0.10 at 5.2 Mb was found across several chromosomes. Seventeen significant (P ≤ 8.17 × 10-5 (-log10 (p) = 4.088)) MTAs of interest, including six that pertained to yield-related traits, were identified using TASSEL MLM. The latter accounted for 5 to 17% of the phenotypic variation expressed. The highly significant association (P ≤ 8.17 × 10-5) between seed length to width ratio and TcSNP 733 on chromosome 5 was verified with FarmCPU (P ≤ 1.12 × 10-8). Fourteen MTAs were common to both the TASSEL and FarmCPU models at P ≤ 0.003. The most significant yield-related MTAs involved seed number and seed length on chromosome 7 (P ≤ 1.15 × 10-14 and P ≤ 6.75 × 10-05, respectively) and seed number on chromosome 1 (P ≤ 2.38 × 10-05), based on the TASSEL MLM. It was noteworthy that seed length, seed length to width ratio and seed number were associated with markers at different loci, indicating their polygenic nature. Approximately 40 candidate genes that encode embryo and seed development, protein synthesis, carbohydrate transport and lipid biosynthesis and transport were identified in the flanking regions of the significantly associated SNPs and in linkage disequilibrium with them. A significant association of fruit surface anthocyanin intensity co-localised with MYB-related protein 308 on chromosome 4. Testing of a genomic selection approach revealed good predictive value (genomic estimated breeding values (GEBV)) for economic traits such as seed number (GEBV = 0.611), seed length (0.6199), seed width (0.5435), seed length to width ratio (0.5503), seed/cotyledon mass (0.6014) and ovule number (0.6325). The findings of this study could facilitate genomic selection and marker-assisted breeding of cacao thereby expediting improvement in the yield potential of cacao planting material.

Selected bacterial recovery in Trinidadian children with chronic tonsillar disease.

UNLABELLED: Pharyngotonsillitis in children is widely treated with antibiotics. AIM: To examine tonsil surface and core microflora following elective adenotonsillectomy in children. METHODS: Tonsils of 102 Trinidadian children were prospectively examined for surface and core bacteriological culture and identification between 2005-2006. RESULTS: Tonsils (360) yielded 800 isolates of Streptococcus spp. (51.3%), Staphylococcus spp. (42.3%) and Gram-negative genera (6.4%). Surface and core recovery of staphylococci and streptococci were similar (p>0.05). More (p<0.001) surfaces (82.2%) than cores (63.3%) grew Streptococcus spp.; alpha-haemolytic Streptococcus prevalence was higher (p<0.001) than ss-haemolytic Streptococcus on surfaces (74.4% vs. 18.6%) than cores (58.9% vs. 13.7%). Surfaces and cores were not concordant for streptococci (p<0.0004) and alpha-haemolytic Streptococcus (p<0.007). Surface and core ss-haemolytic Streptococcus yield was higher (p<0.05) in 6-16 than 1-5 year olds (31% and 23.8% vs 12.5% and 8%). S. pyogenes surface and core prevalence was (84.6% vs 70%) and (50.0% vs 25.0%) in older and younger children respectively. Klebsiella spp. (6.6 %, 2.2%), Proteus (4.4%, 4.4%) and Pseudomonas (4.4 %, 1.1%) grew on surfaces and cores respectively. CONCLUSION: Tonsil surfaces yield higher surface than core carriage for streptococci overall and for alpha haemolytic streptococci. Older children grow more beta-haemolytic streptococci and are high colonizers of S. pyogenes. Studies probing the mechanisms of streptococcal adhesions in Trinidadian children are suggested.

Microbial Community Structure and Function of Soil Following Ecosystem Conversion from Native Forests to Teak Plantation Forests.

Soil microbial communities can form links between forest trees and functioning of forest soils, yet the impacts of converting diverse native forests to monoculture plantations on soil microbial communities are limited. This study tested the hypothesis that conversion from a diverse native to monoculture ecosystem would be paralleled by a reduction in the diversity of the soil microbial communities. Soils from Teak (Tectona grandis) plantations and adjacent native forest were examined at two locations in Trinidad. Microbial community structure was determined via Illumina sequencing of bacterial 16S rRNA genes and fungal internal transcribed spacer (ITS) regions, and by phospholipid fatty acid (PLFA) analysis. Functional characteristics of microbial communities were assessed by extracellular enzyme activity (EEA). Conversion to Teak plantation had no effect on species richness or evenness of bacterial or fungal communities, and no significant effect on EEA. However, multivariate analyses (nested and two-way crossed analysis of similarity) revealed significant effects (p < 0.05) of forest type (Teak vs. native) upon the composition of the microbial communities as reflected in all three assays of community structure. Univariate analysis of variance identified two bacterial phyla that were significantly more abundant in the native forest soils than in Teak soils (Cyanobacteria, p = 0.0180; Nitrospirae, p = 0.0100) and two more abundant in Teak soils than in native forest (candidate phyla TM7, p = 0.0004; WS6, p = 0.044). Abundance of an unidentified class of arbuscular mycorrhizal fungi (AMF) was significantly greater in Teak soils, notable because Teak is colonized by AMF rather than by ectomycorrihzal fungi that are symbionts of the native forest tree species. In conclusion, microbial diversity indices were not affected in the conversion of native forest to teak plantation, but examination of specific bacterial taxa showed that there were significant differences in community composition.

Relations of microbiome characteristics to edaphic properties of tropical soils from Trinidad.

Understanding how community structure of Bacteria, Archaea, and Fungi varies as a function of edaphic characteristics is key to elucidating associations between soil ecosystem function and the microbiome that sustains it. In this study, non-managed tropical soils were examined that represented a range of edaphic characteristics, and a comprehensive soil microbiome analysis was done by Illumina sequencing of amplicon libraries that targeted Bacteria (universal prokaryotic 16S rRNA gene primers), Archaea (primers selective for archaeal 16S rRNA genes), or Fungi (internal transcribed spacer region). Microbiome diversity decreased in the order: Bacteria > Archaea > Fungi. Bacterial community composition had a strong relationship to edaphic factors while that of Archaea and Fungi was comparatively weak. Bacterial communities were 70-80% alike, while communities of Fungi and Archaea had 40-50% similarity. While each of the three component communities differed in species turnover patterns, soils having relatively similar bacterial communities also housed similar archaeal communities. In contrast, the composition of fungal communities had no correlation to bacterial or archaeal communities. Bacterial and archaeal diversity had significant (negative) correlations to pH, whereas fungal diversity was not correlated to pH. Edaphic characteristics that best explained variation between soils in bacterial community structure were: total carbon, sodium, magnesium, and zinc. For fungi, the best variables were: sodium, magnesium, phosphorus, boron, and C/N. Archaeal communities had two sets of edaphic factors of equal strength, one contained sulfur, sodium, and ammonium-N and the other was composed of clay, potassium, ammonium-N, and nitrate-N. Collectively, the data indicate that Bacteria, Archaea, and Fungi did not closely parallel one another in community structure development, and thus microbiomes in each soil acquired unique identities. This divergence could in part reflect the finding that unknown factor(s) were stronger than edaphic characteristics in shaping fungal and archaeal communities.

Clinical features of a paediatric asthma population in Trinidad.

AIM: Clinical features and management of paediatric asthma, highly prevalent in the Caribbean, was explored in Trinidadian children. MATERIALS: Children (2-16 years) with acute [visiting Accident and Emergency (A&E) in the past 12 months (50.2%, 120)] or stable asthma [controlled during the previous 3 months (49.8%, 119)] were studied. RESULTS: There were more (P < 0.001) boys (71.5%) than girls. Asthma was associated with asthmatic mothers (48.5%), fathers (19.2%) and both parents (15.1%) (P < 0.001). Children's average age of first wheeze was 2.5 (standard deviation = 1.3) years; 30.1% were nebulized before 1 year. In the past 12 months, frequencies of A&E visits were once (20.6%), >1 (61.9%) and >3 times (26.4%) (P < 0.001). Sufferers of exacerbations showed negative logistic regression for age (-0.129, standard error = 0.039, P < 0.001) independent of gender. Acute asthmatics who suffered cough, fever and sore throat in the preceding week were respectively 15.2 [odds ratio (OR) = 15.2, 95% confidence interval (CI) = 6.8-34.0], 13.7 (OR = 13.7, 95% CI = 6.7-28.2) and 3.4 (OR = 3.4, 95% CI = 1.7-6.7) times more likely to suffer wheeze exacerbations than stable children. Most stable children (76.5%) inhaled corticosteroids with salbutamol reliever compared with 22.5% of acute asthmatics, whereas 40.8% of the latter group used salbutamol alone compared with 19.3% of stable children (P < 0.001). CONCLUSION: Childhood wheeze occurs before 3 years and is associated with maternal asthma. Cough, fever and sore throat in the previous week are strongly associated with exacerbations. Defining these associations could enhance preventive approaches to combat childhood asthma.

Achado de bactérias selecionadas em crianças de Trinidad com doença amigdaliana crônica / Selected bacterial recovery in Trinidadian children with chronic tonsillar disease

Faringoamigdalite na população pediátrica é largamente tratada com antibióticos. OBJETIVO: Estudar a microflora presente na superfície e no núcleo de amígdalas após adenoamigdalectomia eletiva em crianças. MÉTODO: Amígdalas de 102 crianças de Trinidad foram prospectivamente estudadas por meio de culturas e identificações bacteriológicas feitas a partir de amostras das superfícies e núcleos de suas amígdalas entre 2005-2006. RESULTADOS: A partir de 360 amígdalas, foram isolados Streptococcus spp. (51,3 por cento), Staphylococcus spp. (42,3 por cento) e Gram-Negativos (6,4 por cento). A identificação de estafilococos e estreptococos tanto na superfície quanto no núcleo foi semelhante (p>0,05). Encontramos mais (p<0,001) Streptococcus spp. nas superfícies (82,2 por cento) do que nos núcleos (63,3 por cento); a prevalência de estreptococos alfa-hemolíticos foi maior (p<0,001) do que aquela de estreptococos beta-hemolíticos nas superfícies (74,4 por cento vs. 18,6 por cento) do que nos núcleos (58,9 por cento vs. 13,7 por cento). Não houve concordância entre superfícies e núcleos com relação a estreptococos (p<0,0004) e estreptococos alfa-hemolíticos (p<0,007). Estreptococos beta-hemolíticos foram mais identificados (p<0,05) em crianças dentre 6-16 anos do que naquelas entre 1-5 anos de idade (31 por cento e 23,8 por cento vs 12,5 por cento e 8 por cento). A prevalência de S. pyogenes na superfície e no núcleo foi de (84,6 por cento vs 70 por cento) e (50,0 por cento vs 25,0 por cento) em crianças de maior faixa etária e crianças mais novas, respectivamente. Klebsiella spp. (6,6 por cento, 2,2 por cento), Proteus (4,4 por cento, 4,4 por cento) e Pseudomonas (4,4 por cento, 1,1 por cento) cresceram nas superfícies e núcleos, respectivamente. CONCLUSÃO: As superfícies amigdalianas tinham mais estreptococos e estreptococos hemolíticos do que seus núcleos. Crianças mais velhas tiveram mais estreptococos beta-hemolíticos, e são altamente colonizadoras...

Pharyngotonsillitis in children is widely treated with antibiotics. AIM: To examine tonsil surface and core microflora following elective adenotonsillectomy in children. METHODS: Tonsils of 102 Trinidadian children were prospectively examined for surface and core bacteriological culture and identification between 2005-2006. RESULTS: Tonsils (360) yielded 800 isolates of Streptococcus spp. (51.3 percent), Staphylococcus spp. (42.3 percent) and Gram-negative genera (6.4 percent). Surface and core recovery of staphylococci and streptococci were similar (p>0.05). More (p<0.001) surfaces (82.2 percent) than cores (63.3 percent) grew Streptococcus spp.; α-haemolytic Streptococcus prevalence was higher (p<0.001) than ß-haemolytic Streptococcus on surfaces (74.4 percent vs. 18.6 percent) than cores (58.9 percent vs. 13.7 percent). Surfaces and cores were not concordant for streptococci (p<0.0004) and α-haemolytic Streptococcus (p<0.007). Surface and core ß-haemolytic Streptococcus yield was higher (p<0.05) in 6-16 than 1-5 year olds (31 percent and 23.8 percent vs 12.5 percent and 8 percent). S. pyogenes surface and core prevalence was (84.6 percent vs 70 percent) and (50.0 percent vs 25.0 percent) in older and younger children respectively. Klebsiella spp. (6.6 percent, 2.2 percent), Proteus (4.4 percent, 4.4 percent) and Pseudomonas (4.4 percent, 1.1 percent) grew on surfaces and cores respectively. CONCLUSION: Tonsil surfaces yield higher surface than core carriage for streptococci overall and for α haemolytic streptococci. Older children grow more β-haemolytic streptococci and are high colonizers of S. pyogenes. Studies probing the mechanisms of streptococcal adhesions in Trinidadian children are suggested.