Environmental Profile of Brazilian Green Coffee (6 pp)

Goal, Scope and Background

Brazil is the world's biggest producer of coffee beans with approx. a 30% market share. Depending on climate conditions, approx. 30 million bags of coffee beans are exported annually from Brazil, while domestic consumption is around 10 million bags, which makes Brazil the world's third largest coffee-consuming country. Therefore, the goal of this paper is to present the LCA of green coffee produced in Brazil for the reference crops 2001/02 and 2002/03 in order to generate detailed production inventory data as well as to identify the potential environmental impacts of its tillage in order to realize how to reduce those impacts and increase the environmental sustainability of this product. Only the inputs and outputs relative to the coffee tillage were considered. The production of fertilizers, correctives and pesticides were not included in the boundary, but only their amounts. The functional unit selected for this study was 1,000 kg of green coffee destined for exportation.

Methods

The LCI was performed according to the ISO 14040 standard series. All information considered in this study (use of water, fossil based energy, fertilizers and chemicals) were taken up in in-depth data collection and evaluation by questionnaires applied on a farm level and/or received by mail. Four Brazilian coffee producer regions were evaluated: Cerrado Mineiro, South of Minas Gerais State, the Marília and Alta Mogiana regions in São Paulo State. These regions have the following geographic coordinates: 44 to 50° W longitude and 18 to 24° S latitude. The data refer to a production of 420,000 coffee bean bags and a productive area of approx. 14,300 ha. The varieties of coffee beans considered in this study were Mundo Novo, Catuaí (yellow and red), Icatu (yellow and red), Catucaí (yellow and red) and Obatã. Farm specific data along with agricultural production data have been combined to elaborate a coffee cultivation inventory, which will be applied in an emissions estimation.

Results and Conclusion

The production of 1,000 kg of green coffee in Brazil requires approx. 11,400 kg of water, 94 kg of diesel, 270 kg of fertilizers as NPK, 900 kg of total fertilizers, 620 kg of correctives, 10 kg of pesticides and 0.05 hectare of annual land use. Outputs related to these functional units are approx. 3,000 kg of waste water from coffee washing, 8,500 kg of waste water from the wet method and 750 kg of organic residue that is reincorporated to the tillage as fertilizer. The publication of an LCI of agricultural products is a fundamental step for understanding the potential environmental impacts of each tillage and then establishes the basis for product sustainability. In this way, this work is the first Brazilian initiative for applying LCA to coffee cultivation.

Recommendation and Perspective

Different agricultural practices demonstrate different environmental profiles. The amount of agricultural pesticide is directly related to agricultural practices as tillage rotation, density of plants, etc. This study supplied important results for a better correlation of the agricultural practices and potential environmental impacts of coffee. Future updates of this study will show the evolution of the natural resource management such as land use, new agricultural practices, lower fertilizers and chemicals use.     

Assessing the environmental profile of orange production in Brazil

Background, aim, and scope

Brazil is the world’s largest orange producing country, with a total planted area of more than 820,000 hectares. The bulk of the oranges produced in Brazil (70%) is processed into frozen concentrated orange juice (FCOJ) by large processing companies. Exports represent around 97% of the total FCOJ produced, making Brazil the largest world producer and exporter of FCOJ. The Brazilian citrus sector accounts for half of the world’s supply of orange juice and for 80% of the juice traded on the international market. The goal of this study is to characterize the Brazilian orange producers in terms of farm size, cultivated varieties, watering system and tillage practices. In addition, the study presents some aspects of the LCI of oranges grown specifically for the production of FCOJ in the two major orange-growing areas in Brazil (the Northern and Southern regions of the State of São Paulo) during reference crop year 2002/03 in order to generate detailed production inventory data and identify the potential environmental impacts of tillage, both of which are key to enable the formulation of sustainability parameters for the production of FCOJ in Brazil.

Methods

This study was performed in compliance with the guidelines and requirements of the ISO 14040 standard series. All information and data considered and evaluated in this study (the use of water, energy, fertilizers, pesticides and soil correctors) were gathered through in-depth questionnaires either filled in directly by the farm manager or completed by the farm manager and sent in by mail. The data cover a total of 367,200 metric tons of oranges produced by 4 million plants in commercial production from a total evaluated area that accounts for 19.5% of the overall production of oranges in the State of São Paulo. The two major orange-producing areas in Brazil located respectively in the Northern and Southern regions of the State of São Paulo were evaluated. Only the inputs and outputs associated with tillage practices and technology were considered in this paper. The environmental aspects of the production of fertilizers and pesticides were not included within the boundaries. The functional unit selected for this study was 1,000 kg of oranges for FCOJ. Farm-specific data have been combined with agricultural production data to construct an orange cultivation model.

Results

The orange varieties for the production of FCOJ evaluated in this study were Pêra, Valência and Natal. The farms investigated had a cultivated area varying from 22 to 7,000 ha, with a plant density ranging from 170 to 500 plants/ha and an average yield of 30,500 kg/ha for mature trees. Depending on the region, the production of 1,000 kg of oranges requires approx. 120 to 4,400 MJ of energy; 0.3 to 36 kg of diesel; 1,100 to 54,500 kg of water for irrigation; 0.3 to 65 kg of fertilizers (NPK); 0.1 to 13.5 kg of pesticides; 8 to 650 kg of soil correctors. The distribution of some inputs across the farms located in the two main Brazilian orange-producing regions is presented, in addition to the total weighted average.

Discussion

In addition to the variations among the regions evaluated, it was generally observed that there were very considerable differences in order of magnitude between the data collected from farms located in the same region. Although the inputs are directly related to the specific characteristics of each farm and the climatic and topographic conditions of its location, this study has identified some farms that are in a good position to reduce the amounts of some inputs and enhance their environmental and economical performance.

Conclusions

Taking into account the aspects evaluated in this study, only 21% of the orange growers showed good performance, i.e., consumption of pesticides, fertilizers and soil correction compounds equal to or lower than the weighted averages, except for “land use”, which was above the average for the majority of these farms. This study provided important results that allow for a better understanding of the agricultural practices involved and the potential environmental impacts of this product.

Recommendation and perspective

The authors would like to suggest that the five best-performing farms average values be set as the maximum allowable level and point of departure for the discussion on the good environmental performance of Brazilian FCOJ. Besides the total inputs, the role of good agricultural practices to achieve good yields and, at the same time, improve the sustainability of orange growing should also be taken into consideration. Future updates of this study will show the evolution of natural resource management as a result of improved land use, new agricultural practices, reduced use of fertilizers and agrochemicals.     

Phytotoxicity of pesticides mancozeb and chlorpyrifos: correlation with the antioxidative defence system in <Emphasis Type="Italic">Allium cepa</Emphasis>

Pesticides are a group of chemical substances which are widely used to improve agricultural production. However, these substances could be persistent in soil and water, accumulative in sediment or bio-accumulative in biota depending on their solubility, leading to different types of environmental pollution. The present study was done to assess the impact of pesticides-mancozeb and chlorpyrifos, via morphological and physiological parameters using Allium cepa test system. Phytotoxic effects of pesticides were examined via germination percentage, survival percentage, root and shoot length, root shoot length ratio, seedling vigor index, percentage of phytotoxicity and tolerance index. Oxidative stress on Allium seedlings caused by pesticides was also assessed by investigating the activity of antioxidative enzymes viz. catalase, peroxidase and superoxide dismutase. Correlation was worked out between morphological parameters and antioxidative enzymes to bring out the alliance between them. Mancozeb and chlorpyrifos concentrations were significantly and positively correlated with the activity of antioxidative enzymes and negatively correlated with morphological parameters. Significant positive correlation between various morphological parameters showed their interdependency. However, negative correlation was obtained between activity of antioxidative enzymes and morphological parameters. The enzymes however, showed positive correlation with each other. Based on our result we can conclude that all morphological parameters were adversely affected by the two pesticides as reflected by phytotoxicity in Allium. Their negative correlation with activity of antioxidative enzymes indicates that upregulation of antioxidative enzymes is not sufficient to overcome the toxic effect, thereby signifying the threat being caused by the regular use of these pesticides.     

Sublethal pesticide exposure disrupts courtship in the striped lynx spider,Oxyopes salticus (Araneae: Oxyopidae)

The striped lynx spider, Oxyopes salticus, is a dominant predator of pests in agricultural crops where pesticides are readily applied. While past research shows pesticides directly affect the mortality of agricultural pest predators, recent research has found that pesticide exposure may cause sublethal effects in these organisms. This study examined how residues of three common pesticides (active ingredients: bifenthrin, carbaryl, malathion) impact courtship behaviour of O. salticus. Males provided with cues left by females readily performed courting behaviours. However, when females left cues on a substrate containing bifenthrin and carbaryl, males were less likely to perform courtship behaviours and did so for less time. While males on the malathion treatment showed a trend for reduced courting, this was not significantly different from the treatment without pesticides. The cue produced by females is unknown in this species. However, our study found males reduced their courting over time, suggesting that the cue is ephemeral and not long lived. This study adds important insight into the growing evidence that pesticides greatly impact the ecology of agricultural pest predators and other non‐target organisms by altering their behaviour.     

Development of an Enzyme Linked Immunosorbent Assay and an Immunochromatographic Assay for Detection of Organophosphorus Pesticides in Different Agricultural Products

Objective

Organophosphorus (OP) pesticides are considered hazardous substances because of their high toxicity to nontarget species and their persistence in the environment and agricultural products. Therefore, it is important to develop a rapid, sensitive, and economical method for detecting OP pesticides and their residues in food and the environment.

Methods

A broad, selective monoclonal antibody (MAb) for organophosphorus pesticides was produced. Based on the MAb, an enzyme linked immunosorbent assay (ELISA) and an immunochromatography assay (ICA) for detecting OP pesticides in different agricultural products were developed using a binding inhibition format on microtiter plates and a membrane strip, respectively.

Results

Under the optimized conditions, the IC₅₀ values of the ELISA ranged from 3.7 to 162.2 ng mL^–1 for the 8 OP pesticides. The matrix interferences of Apple, Chinese cabbage, and greengrocery were removed by 40-fold dilution, the recoveries from spiked samples ranged from 79.1% to 118.1%. The IC₅₀ values of ICA for the 8 OP pesticides ranged from 11.8 to 470.4 ng mL⁻¹. The matrix interference was removed from the Chinese cabbage and Apple samples with 5-fold dilution, and the interference was removed from the greengrocery samples with 20-fold dilution. The recoveries from the spiked samples ranged between 70.6 and 131.9%. The established ELISA and ICA were specific selectivity for the 8 OP pesticides.

Conclusions

The established ELISA is a sensitive screening method for the detection of OP pesticides, but the ELISA detection method depends on a laboratory platform and requires a relative long assay time and several steps operation. The established ICA is very useful as a screening method for the quantitative, semi-quantitative or qualitative detection of OP pesticides in agricultural products, and it has advantages over ELISA methods with regard to factors such as the testing procedure, testing time, and matrix interferences, among others.     

Py-GC/MS analysis of sediments from Lake Biwa,Japan: characterization and sources of humic acids

Humic substances extracted and purified from bottom sediments of northern Lake Biwa, Japan, in November 2012 and 2013 were characterized using elemental analysis, Fourier-transform infrared spectroscopy, hydrogen-1 nuclear magnetic resonance (¹H NMR) analysis, and pyrolysis gas chromatography/mass spectrometry (Py-GC/MS). The infrared (IR) bands in the spectra of humic acids showed the presence of amide linkages and polysaccharides. Results of ¹H NMR analysis showed that the humic acids contained approximately twice the number of aliphatic protons as those in the Japanese soil standards used for comparison. Results of the Py-GC/MS analysis, which evaluates pyrolysis temperature dependency of the amount of pyrolysis products, showed that the generation of pyrolysis products in humic acids also differed from that in Japanese soil standards but was similar to that of phytoplankton in Lake Biwa. This analysis method is the first to provide extensive information about the chemical structure of humic substances; conventional Py-GC/MS provides limited information for a single temperature. Data suggest that humic acids in lake sediments are related to chemical characteristics of phytoplankton. Results shed new light on the origins of humic substances in deep-water-lake sediments and provide insights into material recycling in such sediments.     

Sesquiterpene and polyacetylene profile of the Bidens pilosa complex (Asteraceae: Heliantheae) from Southeast of Brazil

The pantropical weed Bidens pilosa (Asteraceae) is a species with several taxonomic problems. Recently it has been shown to be a complex of different species. To shed light on this problem in Brazil, dichloromethane extract of leaves of several populations corresponding to three proposed species for the complex in southeastern Brazil (B. pilosa, Bidens alba and Bidens subalternans) were analyzed by GC–MS. Twenty-four substances were detected, of which four resemble polyacetylenes, the others sesquiterpenes. Five sesquiterpenes tentatively identified as E-caryophyllene, α-humulene, germacrene-D, bicyclogermacrene and α-muurolene were found in all three Bidens species. The polyacetylene phenylhepta-1,3,5-triyne was identified only in B. alba. Multivariate analysis (cluster and principal component analyses) separated the three entities, suggesting that these compounds could represent a useful tool to distinguish species in the B. pilosa complex.     

Macroinvertebrate/sediment relationships along a pesticide gradient in Danish streams

Twenty-nine Danish streams were investigated in September 1998. The streams drained catchments of varying hydrology, topography, soil type and land use. In each stream, the newly accumulated streambed sediment was sampled and subsequently analysed for pesticides. In each stream, five replicate macroinvertebrate samples were taken in the same sediments as the pesticide samples. In addition, five samples were taken in riffles to provide a complete picture of macroinvertebrate community composition. The number of detected pesticides reflected soil type and land use: in agricultural catchments on clay soils the average number of pesticides were 4.3±2.2; on sandy soils 2.6±1.0, while only 1.5±0.6 pesticides were found in streams without agricultural activities. The macroinvertebrate composition showed clear changes along this gradient of sediment pesticide concentrations. The number of the amphipod Gammarus pulex was negatively correlated with sediment pesticide concentration, while the number of Oligochaeta and Hirudinae was positively correlated. The findings indicate that pesticides have a potential to structure macroinvertebrate communities in Danish streams. However, agricultural impact is more than pesticides, and several other factors, such as channelisation, affect the macroinvertebrate community and these are not easily separated.     

Estimating 20‐year land‐use change and derived CO2 emissions associated with crops,pasture and forestry in Brazil and each of its 27 states

Land‐use change (LUC) in Brazil has important implications on global climate change, ecosystem services and biodiversity, and agricultural expansion plays a critical role in this process. Concerns over these issues have led to the need for estimating the magnitude and impacts associated with that, which are increasingly reported in the environmental assessment of products. Currently, there is an extensive debate on which methods are more appropriate for estimating LUC and related emissions and regionalized estimates are lacking for Brazil, which is a world leader in agricultural production (e.g. food, fibres and bioenergy). We developed a method for estimating scenarios of past 20‐year LUC and derived CO₂ emission rates associated with 64 crops, pasture and forestry in Brazil as whole and in each of its 27 states, based on time‐series statistics and in accordance with most used carbon‐footprinting standards. The scenarios adopted provide a range between minimum and maximum rates of CO₂ emissions from LUC according to different possibilities of land‐use transitions, which can have large impacts in the results. Specificities of Brazil, like multiple cropping and highly heterogeneous carbon stocks, are also addressed. The highest CO₂ emission rates are observed in the Amazon biome states and crops with the highest rates are those that have undergone expansion in this region. Some states and crops showing large agricultural areas have low emissions associated, especially in southern and eastern Brazil. Native carbon stocks and time of agricultural expansion are the most decisive factors to the patterns of emissions. Some implications on LUC estimation methods and standards and on agri‐environmental policies are discussed.     

The lethal and sublethal effects of three pesticides on the striped lynx spider (Oxyopes salticus Hentz)

The striped lynx spider, Oxyopes salticus Hentz, is found in high abundances in agricultural fields where it forages on many agricultural pests. Pesticides are applied to these fields and can therefore impact these natural pest predators. Researchers have examined the effects of a number of pesticides on this spider and other pest predators, but many of these studies only examine how these pesticides affect mortality. More recently, researchers have begun to examine the sublethal effects of these chemicals. We examined both the lethal and sublethal effects of three common pesticides with the active ingredients bifenthrin, carbaryl and malathion. We found that only malathion significantly reduced the post‐exposure lifespan of these spiders; however, each pesticide had sublethal effects on behaviour. Exposure to malathion reduced jumping, likely an important foraging and escape behaviour. Spiders exposed to bifenthrin spent increased time grooming, which can reduce the time spent performing other important behaviours. Finally, spiders that were exposed to carbaryl surprisingly increased their prey capture rate. We show here that pesticides can not only directly affect the lifespan of the striped lynx spider but that each pesticide can cause different sublethal effects that likely impact the survival and ecology of these important pest predators.     

Economic Thresholds in Soybean-Integrated Pest Management: Old Concepts, Current Adoption, and Adequacy

Increasing global demands for food underline the need for higher crop yields. The relatively low costs of the most commonly used insecticides in combination with increasing soybean market prices led growers and technical advisors to debate the adequacy of recommended economic thresholds (ETs). The adoption of ETs and pest sampling has diminished in Brazil, leading to excessive pesticide use on soybean. The reduced efficacy of natural biological control, faster pest resurgence, and environment contamination are among the side-effects of pesticide abuse. To address these problems and maximize agricultural production, pest control programs must be guided by a proper integrated pest management (IPM) approach, including the ET concept. Therefore, the most appropriate time to initiate insecticide spraying in soybean is indicated by the available ETs which are supported by experiments over the last 40 years in different edapho-climatic conditions and regions with distinct soybean cultivars. Published scientific data indicate that preventive insecticide use is an expensive and harmful use of chemicals that increases the negative impact of pesticides in agroecosystems. However, the established ETs are for a limited number of species (key pests), and they only address the use of chemicals. There is a lack of information regarding secondary pests and other control strategies in addition to insecticides. It is clear then that much progress is still needed to improve ETs for pest management decisions. Nevertheless, using the current ETs provides a basis for reducing the use of chemicals in agriculture without reducing yields and overall production, thereby improving sustainability.     

Pesticides induced oxidative stress in thymocytes

The role of oxidative stress in immune cell toxicity caused by the pesticides lindane, malathion and permethrin was investigated in thymic cells from C57BL/6 mice. Thymocytes treated with any of these pesticides (concentrations ranging between 50–150 μM) were found to generate both superoxide (^•O₂ ⁻) and H₂O₂. The production of ^•O₂ ⁻ was detected with hydroethidine-ethidium bromide assay. H₂O₂ production was monitored with a flow cytometric fluorescent (DCFH-DA) assay. All three pesticides stimulated ^•O₂ ⁻ release after 5 min exposure. Lindane and permethrin, but not malathion, continued to have significant (p ≤ 0.05) effects on ^•O₂ ⁻ generation following 15 min of exposure. The lindane + malathion mixture was found to cause more-than-additive increase in ^•O₂ ⁻ production compared to individual pesticide treatments (at both 5 and 15 min). However, the effect of the lindane + permethrin mixture was not significantly different than individual components of this mixture. The effects of these pesticides on levels of antioxidant enzymes were also investigated, and only mixtures were found to have significant (p ≤ 0.05) effects. Thus, lindane + malathion and lindane + permethrin mixtures increased total superoxide dismutase (SOD) specific activity, had no effect on catalase levels and inhibited GSH-peroxidase and GSH-reductase specific activities. Although the results of these studies do not explain the mechanism of action of these pesticides on the generation of ^•O₂ ⁻ and H₂O₂, it is worthy of note that mixtures of these chemicals have oxidative responses greater than those of single chemicals. An erratum to this article can be found at     

Occupational exposure of workers to pesticides: Toxicogenetics and susceptibility gene polymorphisms

Farm workers are often exposed to pesticides, which are products belonging to a specific chemical group that affects the health of agricultural workers and is mostly recognized as genotoxic and carcinogenic. The exposure of workers from Piauí, Brazil, to these hazardous chemicals was assessed and cytogenetic alterations were evaluated using the buccal micronucleus assay, hematological and lipid parameters, butyrylcholinesterase (BChE) activity and genetic polymorphisms of enzymes involved in the metabolism of pesticides, such as PON1, as well as of the DNA repair system (OGG1, XRCC1 and XRCC4). Two groups of farm workers exposed to different types of pesticides were evaluated and compared to matched non-exposed control groups. A significant increase was observed in the frequencies of micronuclei, kariorrhexis, karyolysis and binucleated cells in the exposed groups (n = 100) compared to controls (n = 100). No differences were detected regarding the hematological parameters, lipid profile and BChE activity. No significant difference was observed either regarding DNA damage or nuclear fragmentation when specific metabolizing and DNA repair genotypes were investigated in the exposed groups.     

Effects of pesticides on yeasts isolated from agricultural soil

The effect of six various pesticides on the growth of yeasts isolated from agricultural soil was investigated. Two herbicides (with the effective substances lactofen and metazachlor), two fungicides (with the effective substances fluquinconazole and prochloraz), and two insecticides (with the effective substances cypermethrin + chlorpyrifos and triazamate) were tested. It is evident that there are considerable differences in inhibition effects of studied pesticides. The fungicide with the effective substance prochloraz inhibited the growth of majority of yeast strains. The insecticide triazamate at concentration 0.6 mM restricted or inhibited growth of all tested strains. The strains of the genus Cryptococcus were the most sensitive to pesticides, while the strains of the species Cystofilobasidium capitatum, Debaryomyces occidentalis var. occidentalis, and Trichosporon cutaneum were the most resistant.     

Occurrence and distribution of organochlorine pesticide residue levels in water,sediment and aquatic weeds in the Nyando River catchment,Lake Victoria,Kenya

Samples of water, sediments and aquatic weeds were collected from 26 sites in the Nyando River catchment of the Lake Victoria basin in 2005–2006. The objective was to investigate levels of organochlorine pesticides that have either been banned or are restricted for use in Kenya. The pesticides investigated were lindane, aldrin, endosulfan, endrin, dieldrin, DDT, heptachlor and methoxychlor. These pesticides had previously found wide applications in public health and agriculture in Kenya for control of disease vectors and crop pests respectively. Results showed that mean concentrations were highest for methoxychlor (8.817 ± 0.020?µg l^?1) in water, sediments (92.893 ± 3.039 µg kg^?1), and weeds (39.641 ± 3.045?µg kg^?1), the weeds also tended to accumulate aldrin (15.519 ± 3.756?µg kg^?1). The results show that the pesticides are still in use and are detected in the catchment. Stringent management and public awareness measures are required to enforce the ban on the organochlorine pesticides in order to safeguard the environment and ecosystems of Lake Victoria.     

Patterns of land use,extensification, and intensification of Brazilian agriculture

Sustainable intensification of agriculture is one of the main strategies to provide global food security. However, its implementation raises enormous political, technological, and social challenges. Meeting these challenges will require, among other things, accurate information on the spatial and temporal patterns of agricultural land use and yield. Here, we investigate historical patterns of agricultural land use (1940–2012) and productivity (1990–2012) in Brazil using a new high‐resolution (approximately 1 km²) spatially explicit reconstruction. Although Brazilian agriculture has been historically known for its extensification over natural vegetation (Amazon and Cerrado), data from recent years indicate that extensification has slowed down and was replaced by a strong trend of intensification. Our results provide the first comprehensive historical overview of agricultural land use and productivity in Brazil, providing clear insights to guide future territorial planning, sustainable agriculture, policy, and decision‐making.     

基于生态需水保障的农业生态补偿标准

面向流域农业需水和生态需水间的矛盾问题和协调发展的要求,提出了基于生态需水保障的农业生态补偿标准计算方法。其中考虑农业用水定额计算基于生态需水保障的农业用水短缺,引入水分生产函数模型建立保障生态需水量产生的农业用水短缺与产量损失间的关系,根据不同季节作物产量响应系数的变化,定量确定具有时间和等级差异性的农业生态补偿标准。以保障黄河口生态需水引起的山东引黄灌区农业损失补偿标准分析为实例,计算了冬小麦和夏玉米种植户不同等级的生态补偿标准。结论认为,农业生态补偿标准需根据不同的来水过程及生态需水等级确定,面积稳定和保障功能显著的粮食作物应作为补偿标准计算的依据。     

Investigation of mixotrophic,heterotrophic, and autotrophic growth of Chlorella vulgaris under agricultural waste medium

Growth of Chlorella vulgaris and its lipid production were investigated under autotrophic, heterotrophic, and mixotrophic conditions. Cheap agricultural waste molasses and corn steep liquor from industries were used as carbon and nitrogen sources, respectively. Chlorella vulgaris grew remarkably under this agricultural waste medium, which resulted in a reduction in the final cost of the biodiesel production. Maximum dry weight of 2.62 g L^?1 was obtained in mixotrophic growth with the highest lipid concentration of 0.86 g L^?1. These biomass and lipid concentrations were, respectively, 140% and 170% higher than autotrophic growth and 300% and 1200% higher than heterotrophic growth. In mixotrophic growth, independent or simultaneous occurrence of autotrophic and heterotrophic metabolisms was investigated. The growth of the microalgae was observed to take place first heterotrophically to a minimum substrate concentration with a little fraction in growth under autotrophic metabolism, and then the cells grew more autotrophically. It was found that mixotrophic growth was not a simple combination of heterotrophic and autotrophic growth.     

Pesticide tolerant Azotobacter isolates from paddy growing areas of northern Karnataka, India

A total of 14 Azotobacter strains were isolated from different paddy cultivating soils with pH ranging from 6.5 to 9.5 by using serial dilution agar plate method. The strains were Gram negative, rod shaped, cyst forming and developed brown to black colored colonies, which were glistening, smooth, slimy on Ashby’s agar plates. Biochemically they were positive for biochemical tests namely, indole production, citrate, catalase, carbohydrate fermentation and Voges–Proskauer test. Further, sequence analysis of PCR amplicons obtained from these cultures revealed the presence of five different Azotobacter species viz., Azotobacter vinelandii, Azotobacter salinestris, Azotobacter sp., Azotobacter nigricans subsp. nigricans and Azotobacter tropicalis. Phylogenetically these strains were grouped into two distinct clusters. These strains were tested for their ability to grow on a media containing four different pesticides such as pendimethalin, glyphosate, chloropyrifos and phorate, which are commonly used for the paddy. Out of 14 strains tested, 13 strains were able to grow on a media containing herbicides such as pendimethalin, glyphosate and insecticides like chloropyrifos and phorate. However, five Azotobacter strains were able to grow at higher concentration of 5 % pesticides, without affecting their growth rate. Further, the effect of pesticides on the indole acetic acid (IAA) production by Azotobacter strains was also estimated. Azotobacter-16 strain was found to produce 34.4 μg ml^?l of IAA in a media supplemented with 1,000 mg of tryptophan and 5 % of pendimethalin. Present study reveals that species of Azotobacter are able to grow and survive in the presence of pesticides and no significant effects were observed on the metabolic activities of Azotobacter species.