Multiplex PCR method for the detection of human norovirus, Salmonella spp., Shigella spp., and shiga toxin producing Escherichia coli in blackberry, coriander, lettuce and strawberry.

A multiplex PCR method was developed for the simultaneous detection of murine norovirus (MNV-1) as a surrogate for human norovirus (HuNoV) GI and GII, Salmonella spp., Shigella spp., and Shiga toxin producing Escherichia coli (STEC) in fresh produce. The toxicity of the glycine buffer on bacterial pathogens viability was evaluated. The growth of each of the three pathogens (previously stressed) was evaluated at 35 and 41.5 °C in modified buffered peptone water (mBPW) and trypticase soy broth (TSB), supplemented with vancomycin, novobiocin and brilliant green at two concentration levels. The selected conditions for simultaneous enrichment were: 41.5 °C/mBPW/supplemented with 8 ppm vancomycin, 0.6 ppm novobiocin and 0.2 ppm brilliant green. The pathogens and aerobic plate count (APC) growth was evaluated in the enrichment of lettuce, coriander, strawberry and blackberry under the best enrichment conditions. Starting from 1 to 10 CFU/mL, Salmonella reached from 7.63 to 8.91, Shigella 6.81 to 7.76 and STEC 7.43 to 9.27 log CFU/mL. The population reached for the APC was 5.11-6.56 log CFU/mL. Simultaneous detection by PCR was done using designed primers targeting invA, ipaH, stx1 and stx2 genes, and MNV-1. The detection sensitivity was 10-100 PFU for the MNV-1 and 1-10 CFU for each pathogenic bacteria. This protocol takes 6 h for MNV-1 and 24 h for Salmonella spp., Shigella spp., and STEC detection from the same food portion. In total, 200 samples were analyzed from retail markets from Queretaro, Mexico. Two strawberry samples were positive for HuNoV GI and one lettuce sample was positive for STEC. In conclusion, the method developed in this study is capable of detecting HuNoV GI and GII, Salmonella spp., Shigella spp and STEC from the same fresh produce sample.

Flavobacterium agri sp. nov., a novel bacterial species isolated from rhizospheric soil of Coriandrum sativum.

A Gram-stain-negative, strictly aerobic, cream yellow colored, non-motile, rod-shaped bacterial strain, designated MAH-1T was isolated from rhizospheric soil of Coriandrum sativum. A polyphasic taxonomic study was performed on the isolated strain. Optimal growth occurred at 28-30 °C, pH 6.5 and 0% NaCl. The strain showed activity for both catalase and oxidase tests. Cell growth occurs on R2A agar, nutrient agar and Luria-Bertani agar. Cells were able to hydrolyze starch, aesculin, gelatin, and Tween 20. Alignment of 16S rRNA gene sequences indicated that strain MAH-1T was associated with the genus Flavobacterium and was most closely related to Flavobacterium longum YIT 12745T (94.5% sequence similarity) and Flavobacterium caeni LM5T (93.0%). Strain MAH-1T had a genome size of 3,975,600 bp. Genome contained 67 contigs encoded by 3,522 protein-coding genes with 38 tRNA and 6 rRNA genes. The genomic DNA G + C contents of strain MAH-1T was 47.1 mol %. The genomic ANI and dDDH values between strain MAH-1T and one of the close relatives F. caeni LM5T were 72.2 and 18.8%, respectively. The major fatty acids were C15:0 iso, C16:0 iso and C15:0 anteiso. The predominant respiratory quinone was menaquinone 6 (MK-6). Based on physiological, biochemical and phylogenetic data for this isolate, it was confirmed that strain MAH-1T was affiliated to the genus Flavobacterium and represented a novel species, for which the name Flavobacterium agri sp. nov. is proposed. The type strain is MAH-1T (= KACC 19300T = CGMCC 1.16617T).

Hindering of Cariogenic Streptococcus mutans Biofilm by Fatty Acid Array Derived from an Endophytic Arthrographis kalrae Strain.

Streptococcus mutans has been considered as the major etiological agent of dental caries, mostly due to its arsenal of virulence factors, including strong biofilm formation, exopolysaccharides production, and high acid production. Here, we present the antivirulence activity of fatty acids derived from the endophytic fungus Arthrographis kalrae isolated from Coriandrum sativum against Streptococcus mutans. The chemical composition of the fatty acids was analyzed by gas chromatography-mass spectrometry GC-MS and revealed nine compounds representing 99.6% of fatty acids, where unsaturated and saturated fatty acids formed 93.8% and 5.8 % respectively. Oleic and linoleic acids were the major unsaturated fatty acids. Noteworthy, the fatty acids at the concentration of 31.3 mg L-1 completely inhibited Streptococcus mutans biofilm, and water insoluble extracellular polysaccharide production in both polystyrene plates, and tooth model assay using saliva-coated hydroxyapatite discs. Inhibition of biofilm correlated significantly and positively with the inhibition of water insoluble extracellular polysaccharide (R=1, p <0.0001). Furthermore, Arthrographis kalrae fatty acids at a concentration of 7.8 mg L-1 exhibited acidogenesis-mitigation activity. They did not show bactericidal activity against Streptococcus mutans and cytotoxic activity against human oral fibroblast cells at the concentration used. On the other hand, saliva-coated hydroxyapatite discs treated with sub-minimum biofilm inhibitory concentration of fatty acids showed disturbed biofilm architecture with a few unequally distributed clumped matrices using fluorescence microscopy. Our findings revealed that the intracellular fatty acid arrays derived from endophytic Arthrographis kalrae could contribute to the biofilm-preventing alternatives, specifically Streptococcus mutans biofilms.

Occurrence and diversity of ciliated protozoa in the most commonly consumed leafy greens in Brazil: a qualitative study / Ocorrência e diversidade de protozoários ciliados nos vegetais folhosos mais consumidos no Brasil: um estudo qualitativo

Ciliated protozoa are an ubiquitous group of eukaryotes that have been poorly reported on leafy greens. The present study aimed to verify the occurrence and diversity of ciliated protozoa in the leaves and roots of three of the most commonly consumed leafy greens in Brazil – lettuce, rocket and coriander. The vegetable samples were washed by manual agitation (3 minutes) in two different media (mineral water and Page ́s Amoeba Saline solution). After washing, the contents were incubated in Petri dishes and aliquots were removed for microscopic identification and in vivo observation. A total of 21 ciliated protozoa species were found, most of which were bacterivorous. Leafy greens have commonly been associated with foodborne outbreaks and ciliated protozoa, which although they are not a Public Health concern, can act as “Trojan Horses” harboring bacteria, viruses and other protozoa cysts and oocysts and can suggest a new route towards microbiological quality related to the food chain. This is the first report of ciliated protozoa on leafy greens consumed in Brazil.

Os protozoários ciliados constituem um grupo onipresente de eucariotos pouco relatados em vegetais folhosos. O presente estudo teve como objetivo verificar a ocorrência e a diversidade de protozoários ciliados nas folhas e raízes de três dos vegetais folhosos mais consumidos no Brasil - alface, rúcula e coentro. As amostras de vegetais foram lavadas por agitação manual (3 minutos) em dois meios diferentes (água mineral e Solução Salina para Ameba). Após a lavagem, o conteúdo foi incubado em placas de Petri e alíquotas foram retiradas para identificação microscópica e observação in vivo dos organismos. Um total de 21 espécies de protozoários ciliados foi encontrado, a maioria das quais era bacterívora. Tais hortaliças têm sido comumente associadas a surtos transmitidos por alimentos e os protozoários ciliados que, embora não sejam um problema de saúde pública, podem atuar como “Cavalos de Tróia” ao abrigar bactérias, vírus além de cistos e oocistos de protozoários o que pode sugerir uma nova rota para a avaliação da qualidade microbiológica relacionada à cadeia alimentar. Este é o primeiro relato de protozoários ciliados em folhas verdes consumidas no Brasil.

Exploitation of nitric oxide donors to control bacterial adhesion on ready-to-eat vegetables and dispersal of pathogenic biofilm from polypropylene.

BACKGROUND: Nitric oxide (NO) donors have been used to control biofilm formation. Nitric oxide can be delivered in situ using organic carriers and acts as a signaling molecule. Cells exposed to NO shift from biofilm to the planktonic state and are better exposed to the action of disinfectants. In this study, we investigate the capability of the NO donors molsidomine, MAHAMA NONOate, NO-aspirin and diethylamine NONOate to act as anti-adhesion agents on ready-to-eat vegetables, as well as dispersants for a number of pathogenic biofilms on plastic. RESULTS: Our results showed that 10 pM molsidomine reduced the attachment of Salmonella enterica sv Typhimurium 14 028 to pea shoots and coriander leaves of about 0.5 Log(CFU/leaf) when compared with untreated control. The association of 10 pmol L-1 molsidomine with 0.006% H2 O2 showed a synergistic effect, leading to a significant reduction in cell collection on the surface of the vegetable of about 1 Log(CFU/leaf). Similar results were obtained for MAHMA NONOate. We also showed that the association of diethylamine NONOate at 10 mmol L-1 and 10 pmol L-1 with the quaternary ammonium compound diquat bromide improved the effectiveness of biofilm dispersal by 50% when compared with the donor alone. CONCLUSIONS: Our findings reveal a dual role of NO compounds in biofilm control. Molsidomine, MAHMA NONOate, and diethylamine NONOate are good candidates for either preventing biofilm formation or dispersing biofilm, especially when used in conjunction with disinfectants. Nitric oxide compounds have the potential to be developed into a toolkit for pro-active practices for good agricultural practices (GAPs), hazard analysis and critical control points (HACCP), and cleaning-in-place (CIP) protocols in industrial settings where washing is routinely applied. © 2020 Society of Chemical Industry.

Increase in phenolic compounds of Coriandrum sativum L. after the application of a Bacillus halotolerans biofertilizer.

BACKGROUND: There is an urgent need for a new sustainable way of satisfying the increasing demand for food worldwide. One of the main challenges is replacing chemical fertilizers with biofertilizers, which include plant root-associated beneficial microorganisms. The present study reports, for the first time, the effects of SCCPVE07 bacterial strain with respect to improving not only plant development, but also the nutritional content and bioactive compounds content of Coriandrum sativum L., one of the most economically important crops, even for plant growth under salinity stress. RESULTS: Innoculated coriander plants (C. sativum L.) showed an increase in potassium, carbon, calcium and iron content. A significant improvement in phenolic compounds contents was also observed. The contents of 5-O-caffeoylquinic acid, cinnamic acid, 4-methoxy-cinnamic acid hexoside, K-3-O rutinoside, Q-3-O-rutinoside, Q-3-O-glucoside and Q-3-O-glucuronide were significantly enhanced. Moreover, an efficient bacterial root colonization and a noted growth promotion were demonstrated. Bacterial genome was sequenced and analysed. Gene coding related to Plant growth promotion (PGP) mechanisms and proteins involved in plant defence from salinity or in the metabolism of phenolic compounds, such as quercetin 2,3-dioxygenase and phenolic acid decarboxylase, were identified. CONCLUSION: The results obtained in the present study show, for the first time, the beneficial effects of the inoculation of a bacterial Bacillus halotolerans biofertilizer on coriander crops with respect to increasing the content in bioactive compounds and plant development. © 2020 Society of Chemical Industry.

Does nano silver promote the selection of antibiotic resistance genes in soil and plant?

Growing evidences have demonstrated that heavy metal contamination can promote the proliferation of antibiotic resistance genes (ARGs) via co-selection. However, effects of nano-metal-materials on the occurrence and level of ARGs in the soil and plant, have not been fully explored. To gain insights into this impact, we conducted a pot experiment by adding nano­silver particles (AgNPs) as a stimuli and Ag ion (AgNO3) and tetracycline as a comparison. By using high throughput quantitative PCR, our results indicated that application of AgNPs (~20 nm and ~50 nm) at a concentration of 100 ppm resulted in no significant changes in the abundance of ARGs in either soil or phyllosphere (P > 0.05). Nevertheless, the overall pattern of resistome, especially in soil, was shifted following AgNPs application, with a significance increase in the relative abundance of efflux pumps genes, which is an important mechanism for co-selection of ARGs by heavy metals. By comparison, Ag ion at an equivalent Ag mass of AgNPs markedly increased ARGs abundance and shifted ARGs profile in soil, indicating that free Ag ion had a stronger impact on ARGs than AgNPs. These findings provide new insights in assessing the risks of manufactured nanomaterials accumulated in the environment.

Transcriptome profiling of coriander: a dual purpose crop unravels stem gall resistance genes.

Stem gall (Protomyces macrosporus Unger), a serious disease that affects leaves, petioles, stems and fruits of coriander (Coriandrum sativum L.) causing heavy loss in yield. Genetic improvement of coriander for stem gall disease is indispensable. Coriander cultivars of stem gall resistance (ACr-1) and susceptible (CS-6) leaf samples were utilized and transcriptome sequenced using Illumina NextSeq500 platform. After trimming low-quality reads and adapter sequences, a total of 49,163,108 and 43,746,120 high-quality reads were retained and further assembly resulted validated transcripts of 59,933 and 56,861. We have predicted 52,506 and 48,858 coding sequences (CDS) ofwhich 50,506 and 46,945 were annotated using NCBInr database. Gene ontology analysis annotated 19,099 and 17,625 terms; pathway analysis obtained 24 different functional pathway categories; signal transduction, transport, catabolism, translation and carbohydrate metabolism pathways etc. were dominated. Differentially expressed genes analysis predicted 13,123 CDS commonly expressed of which 431 and 400 genes were significantly upregulated and downregulated, respectively, in which Rgenes, stress inducible transcription factors such as ERF, NAC, bZIP, MYB, DREB and WRKY and antifungal related genes were predicted. The real-time PCR analysis of HSP20 gene expression in resistance showed upregulation by 10-fold over susceptible sample and 18s used as a housekeeping gene for normalization. The present results provide an insights into various aspects underlying the development of resistance to stem gall in coriander.

The Transferable Resistome of Produce.

Produce is increasingly recognized as a reservoir of human pathogens and transferable antibiotic resistance genes. This study aimed to explore methods to characterize the transferable resistome of bacteria associated with produce. Mixed salad, arugula, and cilantro purchased from supermarkets in Germany were analyzed by means of cultivation- and DNA-based methods. Before and after a nonselective enrichment step, tetracycline (TET)-resistant Escherichia coli were isolated and plasmids conferring TET resistance were captured by exogenous plasmid isolation. TET-resistant E. coli isolates, transconjugants, and total community DNA (TC-DNA) from the microbial fraction detached from leaves or after enrichment were analyzed for the presence of resistance genes, class 1 integrons, and various plasmids by real-time PCR and PCR-Southern blot hybridization. Real-time PCR primers were developed for IncI and IncF plasmids. TET-resistant E. coli isolated from arugula and cilantro carried IncF, IncI1, IncN, IncHI1, IncU, and IncX1 plasmids. Three isolates from cilantro were positive for IncN plasmids and blaCTX-M-1 From mixed salad and cilantro, IncF, IncI1, and IncP-1ß plasmids were captured exogenously. Importantly, whereas direct detection of IncI and IncF plasmids in TC-DNA failed, these plasmids became detectable in DNA extracted from enrichment cultures. This confirms that cultivation-independent DNA-based methods are not always sufficiently sensitive to detect the transferable resistome in the rare microbiome. In summary, this study showed that an impressive diversity of self-transmissible multiple resistance plasmids was detected in bacteria associated with produce that is consumed raw, and exogenous capturing into E. coli suggests that they could transfer to gut bacteria as well.IMPORTANCE Produce is one of the most popular food commodities. Unfortunately, leafy greens can be a reservoir of transferable antibiotic resistance genes. We found that IncF and IncI plasmids were the most prevalent plasmid types in E. coli isolates from produce. This study highlights the importance of the rare microbiome associated with produce as a source of antibiotic resistance genes that might escape cultivation-independent detection, yet may be transferred to human pathogens or commensals.

Quantitative microbial risk assessment to estimate the risk of diarrheal diseases from fresh produce consumption in India.

This study estimates illness (diarrhea) risks from fecal pathogens that can be transmitted via fecal-contaminated fresh produce. To do this, a quantitative microbial risk assessment (QMRA) framework was developed in National Capital Region, India based on bacterial indicator and pathogen data from fresh produce wash samples collected at local markets. Produce wash samples were analyzed for fecal indicator bacteria (Escherichia coli, total Bacteroidales) and pathogens (Salmonella, Shiga-toxin producing E. coli (STEC), enterohemorrhagic E. coli (EHEC)). Based on the E. coli data and on literature values for Cryptosporidium and norovirus, the annual mean diarrhea risk posed by ingestion of fresh produce ranged from 18% in cucumbers to 59% in cilantro for E. coli O157:H7, and was <0.0001% for Cryptosporidium; for norovirus the risk was 11% for cucumbers and up to 46% for cilantro. The risks were drastically reduced, from 59% to 4% for E. coli O157:H7, and from 46% to 2% for norovirus for cilantro in post-harvest washing and disinfection scenario. The present QMRA study revealed the potential hazards of eating raw produce and how post-harvest practices can reduce the risk of illness. The results may lead to better food safety surveillance systems and use of hygienic practices pre- and post-harvest.

Transfer and Redistribution of Salmonella Typhimurium LT2 and Escherichia coli O157:H7 during Pilot-Scale Processing of Baby Spinach, Cilantro, and Romaine Lettuce.

Several outbreaks of foodborne illness traced to leafy greens and culinary herbs have been hypothesized to involve cross-contamination during washing and processing. This study aimed to assess the redistribution of Salmonella Typhimurium LT2 during pilot-scale production of baby spinach and cilantro and redistribution of Escherichia coli O157:H7 during pilot-scale production of romaine lettuce. Four inoculated surrogate:uninoculated product weight ratios (10:100, 5:100, 1:100, and 0.5:100) and three inoculation levels (103, 101, and 10-1 CFU/g) were used for the three commodities. For each of three trials per condition, 5-kg batches containing uninoculated product and spot-inoculated surrogate products at each ratio and inoculation level were washed for 90 s in a 3.6-m-long flume tank through which 890 L of sanitizer-free, filtered tap water was circulated. After washing and removing the inoculated surrogate products, washed product (∼23, 225-g samples per trial) was analyzed for presence or absence of Salmonella Typhimurium or E. coli O157:H7 by using the GeneQuence Assay. For baby spinach, cilantro, and romaine lettuce, no significant differences ( P > 0.05) in the percentage of positive samples were observed at the same inoculation level and inoculated:uninoculated weight ratio. For each pathogen product evaluated (triplicate trials), inoculation level had a significant impact on the percentage of positive samples after processing, with the percentage of positive samples decreasing, as the initial surrogate inoculation level decreased. The weight ratio of contaminated:noncontaminated product plays an important role: positive samples ranged from 0% to 11.6% ± 2.05% and from 68.1% ± 33.6% to 100% among the four ratios at inoculation of 10-1 and 101 CFU/g, respectively. To our knowledge, this study is the first to assess the redistribution of low levels of pathogens from incoming product to leafy greens during processing and should provide important data for microbial risk assessments and other types of food safety analyses related to fresh-cut leafy greens.

Potassium permanganate cleansing is an effective sanitary method for the reduction of bacterial bioload on raw Coriandrum sativum.

OBJECTIVE: Raw vegetables including flowers, leaves, stems, and roots are important carriers of food borne pathogens. We evaluated the bacteriological contamination of unwashed coriander leaves, and effectiveness of cleansing with 0.1% potassium permanganate solution as decontamination method. RESULTS: Significant bacterial contamination including pathogens like Salmonella species and Aeromonas species were isolated from unwashed coriander leaves. Decontamination with 0.1% potassium permanganate was found to be more effective than three steps wash with sterile water.

Improvement in shelf life of minimally processed cilantro leaves through integration of kinetin pretreatment and packaging interventions: Studies on microbial population dynamics, biochemical characteristics and flavour retention.

Effect of integrating optimized combination of pretreatment with packaging on shelf life of minimally processed cilantro leaves (MPCL) was appraised through analysis of their sensory attributes, biochemical characteristics, microbial population and flavour profile during storage. Minimally pretreated cilantro leaves pretreated with 50ppm kinetin and packed in 25µ polypropylene bags showed a shelf life of 21days. Optimized combination helped in efficiently maintaining sensory parameters, flavour profile, and retention of antioxidants in MPCL until 21days. Studies conducted on the effect of optimized combination on microbial population and flavour profile revealed that among different microorganisms, pectinolysers had a significant effect on spoilage of MPCL and their population of ⩽3.59logcfu/g was found to be acceptable. Principal component analysis of headspace volatiles revealed that (E)-2-undecenal, (E)-2-hexadecenal, (E)-2-tetradecenal & (E)-2-tetradecen-1-ol in stored samples clustered with fresh samples and therefore, could be considered as freshness indicators for MPCL.

Prevalence and behavior of multidrug-resistant shiga toxin-producing Escherichia coli, enteropathogenic E. coli and enterotoxigenic E. coli on coriander.

The prevalence and behavior of multidrug-resistant diarrheagenic Escherichia coli pathotypes on coriander was determined. One hundred coriander samples were collected from markets. Generic E. coli were determined using the most probable number procedure. Diarrheagenic E. coli pathotypes (DEPs) were identified using two multiplex polymerase chain reaction procedures. Susceptibility to sixteen antibiotics was tested for the isolated DEPs strains by standard test. The behavior of multidrug-resistant DEPs isolated from coriander was determined on coriander leaves and chopped coriander at 25°± 2 °C and 3°± 2 °C. Generic E. coli and DEPs were identified, respectively, in 43 and 7% of samples. Nine DEPs strains were isolated from positive coriander samples. The identified DEPs included Shiga toxin-producing E. coli (STEC, 4%) enterotoxigenic E. coli (ETEC, 2%) and enteropathogenic E. coli (EPEC, 1%). All isolated DEPs strains exhibited multi-resistance to antibiotics. On inoculated coriander leaves stored at 25°± 2 °C or 3°± 2 °C, no growth was observed for multidrug-resistant DEPs strains. However, multidrug-resistant DEPs strains grew in chopped coriander: after 24 h at 25° ± 2 °C, DEPs strains had grown to approximately 3 log CFU/g. However, at 3°± 2 °C the bacterial growth was inhibited. To the best of our knowledge, this is the first report of the presence and behavior of multidrug-resistant STEC, ETEC and EPEC on coriander and chopped coriander.

Arbuscular mycorrhizal fungi are an alternative to the application of chemical fertilizer in the production of the medicinal and aromatic plant Coriandrum sativum L.

The widespread use of agrochemicals is detrimental to the environment and may exert harmful effects on human health. The consumer demand for organic food plants has been increasing. There is thus a rising need for alternatives to agrochemicals that can foster sustainable plant production. The aim of this study was to evaluate the potential use of an arbuscular mycorrhizal (AM) fungus as an alternative to application of chemical fertilizer for improving growth performance of the medicinal and aromatic plant Coriandrum sativum. Plants were inoculated with the AM fungus Rhizophagus irregularis BEG163 and/or supplemented with a commercial chemical fertilizer (Plant Marvel, Nutriculture Bent Special) in agricultural soil. Plant growth, nutrition, and development of AM fungus were assessed. Plants inoculated with R. irregularis and those supplemented with chemical fertilizer displayed significantly improved growth performances when compared with controls. There were no significant differences in total fresh weight between plants inoculated with R. irregularis or those supplemented with chemical fertilizer. Leaf chlorophyll a + b (82%), shoot nitrogen (44%), phosphorus (254%), and potassium (27%) concentrations increased in plants inoculated with R. irregularis compared to controls. Application of chemical fertilizer inhibited root mycorrhizal colonization and the length of the extraradical mycelium of R. irregularis. Inoculation with R. irregularis was equally or more efficient than application of chemical fertilizer in promoting growth and nutrition of C. sativum. AM fungi may thus contribute to improve biologically based production of food plants and reduce the dependence on agrochemicals in agriculture.

Susceptibility of Klebsiella pneumoniae on coriander leaves to liquid- and vapor-phase ethanol.

The bio-control of ethanol on Klebsiella pneumoniae on fresh coriander leaves for significantly reducing consumer health risk was investigated. Washed and sterilized leaves of coriander were inoculated with K. pneumoniae cultured in Trypticase Soy broth. Susceptibility of the K. pneumoniae to liquid- and evaporated vapor-phase ethanol (EVE) was then examined in vitro Complete inhibition of K. pneumoniae was found with 18% (v/v) liquid ethanol. Exposure for 15 min to EVE (9.00 ± 0.8 mmol L(-1)) completely destroyed K. pneumoniae (4.04 ± 0.02 log CFU/ml) spread on Mueller Hilton agar at 30 ± 2°C. The effect of EVE with and without evaporated water vapor (EWV) on the susceptibility of K. pneumoniae on fresh coriander leaves was examined. While exposure to EVE affected the survival of K. pneumoniae, the degree of reduction depended on both the inoculation level and the EWV. Complete reduction of K. pneumoniae was achieved for the low inoculation level by EVE alone (37 ± 2% relative humidity; RH) but susceptibility was reduced with EWV (high RH; 80 ± 2%). Scanning electron microscope (SEM) images of inoculated coriander leaves confirm the effects of EVE in reducing levels of K. pneumoniae Exposure to EVE alone proved an effective bio-control for K. pneumoniae on fresh coriander leaves.

Arbuscular mycorrhiza differentially affects synthesis of essential oils in coriander and dill.

Research on the role of arbuscular mycorrhizal fungi (AMF) in the synthesis of essential oils (EOs) by aromatic plants has seldom been conducted in field-relevant conditions, and then, only limited spectra of EO constituents have been analyzed. The effect was investigated of inoculation with AMF on the synthesis of a wide range of EO in two aromatic species, coriander (Coriandrum sativum) and dill (Anethum graveolens), in a garden experiment under outdoor conditions. Plants were grown in 4-l pots filled with soil, which was either γ-irradiated (eliminating native AMF) or left non-sterile (containing native AMF), and inoculated or not with an isolate of Rhizophagus irregularis. AMF inoculation significantly stimulated EO synthesis in both plant species. EO synthesis (total EO and several individual constituents) was increased in dill in all mycorrhizal treatments (containing native and/or inoculated AMF) compared to non-mycorrhizal plants. In contrast, EO concentrations in coriander (total EO and most constituents) were increased only in the treatment combining both inoculated and native AMF. A clear positive effect of AMF on EO synthesis was found for both aromatic plants, which was, however, specific for each plant species and modified by the pool of AMF present in the soil.

Cilantro microbiome before and after nonselective pre-enrichment for Salmonella using 16S rRNA and metagenomic sequencing.

BACKGROUND: Salmonella enterica is a common cause of foodborne gastroenteritis in the United States and is associated with outbreaks in fresh produce such as cilantro. Salmonella culture-based detection methods are complex and time consuming, and improvments to increase detection sensitivity will benefit consumers. In this study, we used 16S rRNA sequencing to determine the microbiome of cilantro. We also investigated changes to the microbial community prior to and after a 24-hour nonselective pre-enrichment culture step commonly used by laboratory analysts to resuscitate microorganisms in foods suspected of contamination with pathogens. Cilantro samples were processed for Salmonella detection according to the method in the United States Food and Drug Administration Bacteriological Analytical Manual. Genomic DNA was extracted from culture supernatants prior to and after a 24-hour nonselective pre-enrichment step and 454 pyrosequencing was performed on 16S rRNA amplicon libraries. A database of Enterobacteriaceae 16S rRNA sequences was created, and used to screen the libraries for Salmonella, as some samples were known to be culture positive. Additionally, culture positive cilantro samples were examined for the presence of Salmonella using shotgun metagenomics on the Illumina MiSeq. RESULTS: Time zero uncultured samples had an abundance of Proteobacteria while the 24-hour enriched samples were composed mostly of Gram-positive Firmicutes. Shotgun metagenomic sequencing of Salmonella culture positive cilantro samples revealed variable degrees of Salmonella contamination among the sequenced samples. CONCLUSIONS: Our cilantro study demonstrates the use of high-throughput sequencing to reveal the microbiome of cilantro, and how the microbiome changes during the culture-based protocols employed by food safety laboratories to detect foodborne pathogens. Finding that culturing the cilantro shifts the microbiome to a predominance of Firmicutes suggests that changing our culture-based methods will improve detection sensitivity for foodborne enteric pathogens.

Microbiological analysis of pre-packed sweet basil (Ocimum basilicum) and coriander (Coriandrum sativum) leaves for the presence of Salmonella spp. and Shiga toxin-producing E. coli.

Enteric pathogens, such as Salmonella spp. and pathogenic Escherichia coli, have been detected and associated with food borne outbreaks from (imported) fresh leafy herbs. Screening on imported herbs from South East Asian countries has been described. However, limited information on prevalence of these pathogens is available from other sourcing regions. Therefore, fresh pre-packed basil and coriander leaves from a Belgian trading company were investigated for the presence of Salmonella spp., Shiga toxin-producing E. coli (STEC), generic E. coli and coliforms. In total 592 samples were collected originating from Belgium, Israel and Cyprus during 2013-2014. Multiplex PCR followed by further culture confirmation was used for the detection of Salmonella spp. and STEC, whereas the Petrifilm Select E. coli and VRBL-agar were used, respectively, for the enumeration of E. coli and coliforms. Salmonella was detected in 10 out of 592 samples (25g) (1.7%; 5 from basil and 5 from coriander), of which two samples were sourced from Israel and eight from Cyprus. The presence of STEC was suspected in 11 out of 592 samples (25g) (1.9%; 3 basil and 8 coriander), due to the detection of stx and eae genes, of which one sample originated from Belgium, four from Israel and six from Cyprus. No STEC was isolated by culture techniques, but in three samples a serotype (O26, O103 or O111) with its most likely associated eae-variant (ß or θ) was detected by PCR. Generic E. coli was enumerated in 108 out of 592 samples, whereby 55, 32 and 13 samples respectively between 10-100, 100-1000 and 1000-10,000cfu/g and 8 samples exceeding 10,000cfu/g. Coliforms were enumerated in all herb samples at variable levels ranging from 1.6 to 7.5logcfu/g. Further statistics indicate that the E. coli class (categorized by level) was significantly correlated with the presence of Salmonella (p<0.001) or STEC (p=0.019), while coliform counts were significant correlated with Salmonella (p<0.001), but not with STEC (p=0.405). Generic E. coli class is a better indicator for the presence of enteric pathogens than coliforms on fresh herbs, but the relationship between E. coli and Salmonella or STEC was not strong enough to provide a threshold value for E. coli to assure food safety (i.e. no pathogens present). Results indicate that fresh leafy herbs like basil and coriander sourced from different cultivation regions, may contain enteric pathogens and potentially pose a risk for human health.

Microbial safety and quality of fresh herbs from Los Angeles, Orange County and Seattle farmers' markets.

BACKGROUND: Farmers' markets have been growing in popularity in the United States, but the microbial quality and safety of the food sold at these markets is currently unknown. The purpose of this study was to assess the microbial safety and quality of fresh basil, parsley and cilantro sold at farmers' markets in the Los Angeles, Orange County and greater Seattle areas. RESULTS: A total of 133 samples (52 basil, 41 cilantro and 40 parsley) were collected from 13 different farmers' markets and tested for Salmonella and generic Escherichia coli. One sample (parsley) was confirmed positive for Salmonella and 24.1% of samples were positive for generic E. coli, with a range of 0.70-3.15 log CFU g(-1) . Among the herbs tested, basil showed the highest percentage of samples with generic E. coli (26.9%), followed by cilantro (24.4%) and then parsley (20.0%). For 12% of samples, the levels of generic E. coli exceeded guidelines established by the Public Health Laboratory Service for microbiological quality of ready-to-eat foods. CONCLUSION: Overall, this study indicates the presence of Salmonella and generic E. coli in fresh herbs sold at farmers' markets; however, additional studies are needed to determine the sources and extent of contamination.