The aim of this study was to assess the microbiological profile of leafy vegetables from organic and conventional farming (
The organic food and beverage market sold about 81.6 billion US dollars’ worth of produce worldwide in 2015. Nowadays, the highest per capita consumption of organic food is found in European countries, such as Switzerland, Luxembourg, and Denmark [
The reasons for the consistent growth in the consumption of organic vegetables go from ethical and political reasons to life quality, safety, and environmental concerns [
The relationship between microbiological risk and organic production is still uncertain since many factors may contribute to the development of microorganisms, including differences between the cropping system. The use of animal manure in organic farming naturally raises concern about the microbiological safety. Since there is a restriction concerning the chemicals allowed in organic production [
In some scientific studies greater risk with the consumption of organic food was not identified [
Leafy vegetables do not generally represent the main type of food reported in outbreaks. From 1998 to 2008 only 5.2% of bacterial foodborne illnesses in the USA were related to leafy vegetables [
In this context, the present work proposes to microbiologically evaluate certified organic leafy vegetables and conventional ones from the same cultivars.
Samples of leafy vegetables from organic and conventional cultivation were obtained from two farms located in Cordeiropolis (22°28′55′′S; 47°27′24′′O) and Americana (22°44′21′′S; 47°19′53′′O), both cities in Sao Paulo State, Brazil. The production area for organic cultivation has 12 hectares where 50 different species of vegetables are planted. Well water is used for irrigation which is performed by microirrigation (drip and microspray irrigation). The fertilization is manual and made with manure composting into soils. The vegetables are collected by a work team composed of 8 persons, and each employee is responsible for the collection of a particular plant in order to observe possible changes in the crop. The organic vegetables presented national certification by Ecocert (a system which adopts external audits) and the leafy vegetables that have been evaluated are produced organically there since 2014. The production area for conventional agriculture has 1.5 hectares, in which 19 different species of vegetables are grown throughout the year. The irrigation process is only by sprinkling and the irrigation water comes from a river from to the PCJ watersheds (belonging to Piracicaba, Corumbataí and Jaguari cities). Manure is also used for fertilization using a manual process. The work team is composed by 3 employees. Both producers used the same plant seedlings from the same provider, so the production only differed with respect to the type of cultivation. Five units of each leafy vegetable, organically and conventionally grown, were collected. The following species were analyzed: green leaf lettuce varieties “Milena” (LM) and “Vanda” (LV) and iceberg lettuce (IL) (
Summary of the characteristics of the two evaluated farms.
Location | Fertilization | Water source | Irrigation system | Irrigation system |
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Farm of organic cultivation | Manual with manure composting | Well water (groundwater) | Microirrigation (drip and microspray) | Applies the water through small emitter to the soil surface; drip irrigation allows the water to drip slowly to the roots of plants minimizing the water evaporation; microspray applies water to the soil surface by a small spray or mist |
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Farm of conventional cultivation | Manual with manure | River water (surface water) | Sprinklers | Sprinkler irrigation is a method similar to natural rainfall; water is delivered through a pressurized pipe network to sprinklers which spray the water into the air |
The analyses were performed from February to November 2016. The analysis routine was the following: each type of leafy vegetable obtained by conventional and organic cultivation was harvested at the same day and taken to the Microbiology Laboratory. All analyses were performed on the day the vegetables were harvested. They were placed in plastic bags without modifying the atmosphere and stored at
The samples were analyzed for aerobic mesophilic bacteria, yeasts and molds, coliform (at 30°C and 45°C) counts according to Downes and Ito [
Plate Count Agar (PCA) (Difco/BD, USA) was used for the aerobic mesophilic bacterial count with the addition of 1% triphenyl tetrazolium chloride (TTC) (Merck, USA), using the pour plate technique and incubation at
The coliform counts at 30°C and 45°C were obtained using the Conventional Multiple Tube Technique with Lauryl Sulfate Tryptose Broth (Oxoid, England) and Brilliant Green Bile Broth (Oxoid, England), with incubation at
The method described by Andrews and Hammack [
To identify the species of Enterobacteriaceae present in the samples, the commercial systems Bactray® I and II (Laborclin, Brazil) were used. These consist of a miniature kit indicated for gram-negative oxidase negative bacilli, glucose-fermenting and glucose nonfermenting bacteria. Initially, MacConkey media for the growth of typical colonies was used, with incubation at 35°C/48 h, and then transferred to Brain Heart Infusion broth (BHI) and incubated at 35°C/24 h. The BHI broth was adjusted to a 0.5 McFarland turbidity reading in a spectrophotometer at 600 nm.
An aliquot of 1.0 mL of the suspension was then transferred to the Bactray I and II systems and incubated at 35°C/18 h–24 h. An electronic program provided by Laborclin revealed the species found.
The data obtained were compared with the Brazilian legislation (RDC no. 12) (Resolução da Diretoria Colegiada = Resolution of the Board of Directors) which indicates a maximum count for coliform bacteria at 45°C of
The variables were expressed as the means followed by the standard variation. The distribution histograms, mean, standard deviation, and minimal and maximum values were verified. The Kolmogorov Smirnov test was used to verify the adherence of distributions with the normal curve.
The means obtained for organic and conventional cultivation were compared using Student’s
The comparison between aerobic mesophilic bacteria, coliforms at 30°C and 45°C, and yeasts and molds can be observed in Table
Mean counts of aerobic mesophilic bacteria, yeasts and molds, and coliforms at 30°C and 45°C from 70 vegetable samples from organic and conventional cultivation.
Microorganism | Conventional | Organic |
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Mean | SD± | Mean | SD± | ||
Aerobic mesophilic bacteria ( |
4.84 | 0.83 | 4.32 | 1.13 |
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Yeasts and molds ( |
3.18 | 0.78 | 2.81 | 0.85 |
|
Coliforms at 30°C ( |
1.96 | 0.97 | 1.08 | 1.02 |
|
Coliforms at 45°C ( |
0.79 | 0.88 | 0.25 | 0.27 |
|
SD is standard deviation. Values in bold indicate significant statistical difference (
Counting of aerobic mesophilic bacteria (a), yeasts and molds (b), and coliforms at 30°C (c) and 45°C (d) among the 7 species of organic and conventional vegetables. Equal letters for the same vegetable indicate no statistical difference in Student’s
Of all the samples analyzed, only one conventional vegetable (LV) and two organic ones (LM and CC) presented aerobic mesophilic bacterial counts higher than
A range from 4 to >7
The contamination with coliforms at 30°C found presently reached
The occurrence of coliforms at 45°C can indicate fecal contamination. Lotto and Valarini [
There was a statistical difference between the microbial load (i.e., coliforms at 30°C and at 45°C and yeasts and molds) of the leafy vegetables obtained by the two cultivation systems; however, the absolute numbers of the counts were similar, less than 1 log different. Although the averages were close, in the individual analyses of two conventionally cultivated leafy vegetables (LV (Vanda lettuce) and IL (iceberg lettuce)) higher counts of coliforms were observed at 45°C than the established by the Brazilian legislation [
The presence of
In some Brazilian studies, higher microbial counts were observed for organic vegetables [
The irrigation system is another important factor in the microbiological safety of a crop, since contaminated water can reach the leaves during irrigation [
The indiscriminate use of herbicides and pesticides in conventional agriculture systems has been indicated as one of the causes of water and soil contamination and an imbalance in the environment, with direct impacts on the microbial loads of the soil and on several other species [
Table
Enterobacteriaceae species identified by miniature kits Bactray I and II from samples of organic and conventional cultivation of leafy vegetables.
Vegetable | Enterobacteriaceae | |||
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Conventional | Precision rate (%) | Organic | Precision rate (%) | |
Iceberg lettuce ( |
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Lettuce, var. |
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- |
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49.96 |
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Lettuce, var. |
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84.55 | |
|
99.85 | |||
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Escarole |
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51.83 |
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51.83 |
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Chicory |
|
100 |
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- |
|
100 | |||
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99.2 | |||
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Collard green ( |
|
100 |
|
|
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44.97 | |||
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Parsley |
|
100 |
|
100 |
N.D.: no species detected.
It is also important to point out that, with respect to the species reported in the current study, the reports found in the literature concerning infections caused by them occurred mostly in hospital environments and possibly with immunocompromised subjects. It is also important to highlight that the increase in outbreaks has been strongly associated with the consumption of vegetables by immunocompromised individuals [
Organic agriculture production can be associated with other benefits, especially regarding sustainability and public health. Brazil, for example, is the leading country in the use of pesticides (5.2 L per capita/year), causing a strong impact on the environment in terms of the depletion of natural resources, the persistence of certain residues, and a decrease in biodiversity. The effects on public health due to the consumption of contaminated food, as well as on farm workers due to pesticide intoxication, can be considered as a violation of human rights: violating the right to health and to adequate and healthy food [
The present study showed that the same cultivars of leafy vegetables obtained by organic cultivation had lower microbial counts than those obtained by conventional cultivation.
The methodological approach of the present study does not allow to conclude which cropping system is safer; however, the observed results indicate that leafy vegetables cultivated in a conventional way can present a higher count of microorganisms even after the use of chemosynthetic pesticides and fertilizers. In this sense, good farming practices, independent of the cropping system, tend to be more effective in controlling the microbiological load. The quality of the water and the type of irrigation used in production can affect the quality of the leafy vegetables, so they should be carefully evaluated and implemented.
Anyway, the detection of some Enterobacteriaceae species (but no
The authors declare no conflicts of interest.
The authors are grateful to the National Council for Scientific and Technological Development (CNPq) for the scholarships provided (129891/2019-9 and 303568/2016) and for financial support from the Sao Paulo Research Foundation (FAPESP, Project 2013/04304-4). The authors are grateful also to the students Gisele V. R. Pereira, Larissa T. Silva, and Franciele C. Silva for technical assistance.