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The role of storage on Mancozeb fungicide formulations and their antifungal activity against Fusarium oxysporium and Rhizoctonia solani
-
Received: ,
Accepted: ,
This article was originally published by Elsevier and was migrated to Scientific Scholar after the change of Publisher.
Abstract
The stability of fungicides Dicozeb 80% WP (Mancozeb), Sand flower 72% WP (Cymoxail + Mancozeb) and Triomax 66% WP (Cymoxail + Mancozeb + Copper oxychloride) were tested before and after storage at 54 ± 2 °C for 14 days and investigated their activity on radial growth of F. oxysporum and R. solani in vitro by Poisoned Food Technique. The results indicated that slightly increased of degradation rate of the active ingredient of Triomax followed by Sand flower and Dicozeb. The fungicidal activity of Mancozeb increased when mixed with Cymoxanil and Copper oxychloride by 97.4 and 73.4 times against F. oxysporum and R. solani, respectively. While Cymoxanil increased the activity of Mancozeb by 39.5 and 45.9 times against tested fungi F. oxysporum and R. solani, respectively. In addition, degradation of tested fungicides caused increase the mean activity of polyphenol oxidase enzyme and mean proline amino acid in mycelium of tested fungi. With another point of view, the present of Cymoxanil + Copper oxychloride in mixture of Triomax caused decrease in degradation of fungicidal activity of Mancozeb up on other tested fungicides.
Keywords
Fungicides
Storage
Thermal stability
Ethylenethiourea
Antifungal activity
Enzyme
Amino acid
1 Introduction
Plant diseases play a vital role in the decline of natural resources in agriculture. Their causal agents include biotic and abiotic factors, genetic disorders and living infections agents such as fungi, bacteria, viruses, viroid’s, phytoplasmas, nematodes, parasitic plants, and protozoan’s (Agrios 2005). Approximately 20,000 species of fungal plant pathogens have been report and these fungi (Cooper 2007, Ong 2011) cause about 85% of plant diseases. Many fungal genera including Fusarium, Alternaria, Botrytis, Helminthosporium, Penicillium, and Rhizoctonia have proved as a harmful pathogenic fungi and cause huge loss of crop yield in quality and quantity world-wide (Boyraz & Ozcan 2006).
Fusarium wilt of tomato (Lycopersicon esculentum Mill.) caused by Fusarium oxysporum [(Schlecht.) f. sp. lycopersici (Sacc.)] Snyder et Hansen., is one of the most prevalent, serious diseases of tomato (Reis et al. 2005; Sudhamoy et al., 2009) and the disease is considered as one of the main soil-borne systemic diseases (Schwarz and Grosch 2003). It causes significant losses in tomato production in both greenhouse and field-grown tomates (Nusret and Steven, 2004). Among these diseases, Rhizoctonia black scurf and stem canker caused by the fungus Rhizoctonia solani Kühn, considered as a severe problem in all potato producing zones of the country (Sneh et al., 1991; Ahmad et al., 1995; Khan et al., 1995).
Application of fungicides is the most convenient and predominant way for disease control. Their use has made it feasible to enhance crop yields and food production. The efficacy of fungicides influenced by many biological and environmental factors that directly influence the metabolic activities of fungal cells (Reinprecht 2010; Peerzada et al., 2020).
Dithiocarbamate pesticides form an important class of organic fungicide used over the world to control many diseases in a variety of crops. They are of low acute toxicity, but some of Ethylenebisdithiocarbamate (EBDCS) manb, zinab, mancozeb, metiran and nabam can decompose to give another ethylenthiourea has been shown to possess carcinogenics, mutagenic, goitergenic and tetratogenic activates in animals, (Gronogrorac and schwack, 2009; Larsson et al., 1976).
Mancozeb was one of the compounds in three combinations, which proved best in present finding. Therefore, in these combinations the efficacy may be due to Mancozeb (Bhaliya and Jadeja, 2014).
Ethylenebisdithiocarbamate (EBDCS) fungicide applied to crop to prevent disease such as early blight, late blight and anthracnose, with other diseases such as bacterial speak and bacterial spots controlled by the application of copper salts. Mixed sprays of EBDCS and copper found to give better results bacterial speak than copper alone. EBDCS applied as their manganese and zinc complex Maneb or Mancozeb on the crop. The solubility activity and stability of the dithiocarbamate complex is depend on metal ion (Singh and Tripathi 1982). Combing the Cymoxanil with other fungicides belonging to different mode of action has helped prevent the selection of strains pathogens resistant to this fungicide. Therefore, it is used in two or in three ways mixture with protestants systemic fungicide (Gisi, 1996).
The present study aimed to evaluate the effect of the storage at 54 ± 2 °C for 14 days on fungicides Dicozeb (Mancozeb), Sand flower (Cymoxail + Mancozeb) and Triomax (Cymoxail + Mancozeb + Copper oxychloride) stability as well as their efficacy against the mycelial growth of R. solani and F. oxysporum tested under in vitro condition.
2 Materials and methods
2.1 Chemical fungicides used
Three commercial fungicides used in this study were obtained from the Central Agricultural Pesticides Laboratory, Dokki, Giza, Egypt and tabulated in Table 1.
Common name
Trade Name
Chemical name
Formulation
Manufacturer
Mancozeb
Dicozeb
Manganese ethylenebis ((dithiocarbamate) polymeric) complex with zinc salt.
80% WP
Agrochem
a) Cymoxanil
b) MoncozebSand flower
a)1-(2-cyano-2-metho xymino acetyl)-3-ethylurea.
b) Manganese ethylenebis ((dithiocarbamat) polymeric) complex with zinc salt.72% WP
Xi'an MTI Co.,Ltd
a) Cymoxanil
b) Mancozeb
c) Copper oxychlorideTriomax
a)1-(2-cyano-2-metho xymino acetyl)-3-ethylurea.
b) Manganese sthylenebis ((dithiocarbamate) polymeric) complex with zinc salt.
c) Dicopper chloride trihydroxide.66% WP
Agria S. A.
2.2 Chemicals
Sodium thiosulfate, potassuim iodid, sulfuric acid, nitric acid methanolic potassuim, leadacetate, silver nitrate and all organic solvents, methanol and acetonitrile HPLC grade, Deionized water, Mancozeb 80% WP (C4H6 Mn N2 S4)× (Zn)y under trade name Dicozib, Cymoxanil C7M10N4O3 (4%) + Mancozeb (12%) + Copper oxychloride (50%) equal metallic copper 29% used under trade name Triomax 66% WP and Mancozeb 64% + Cymoxanil 8% used under trade name Sand flower 72% WP.
2.3 Apparatus
An equipment HPLC (Agilent 1200 sires was used DAD detector and C-18 column. The used of a fungicide formulation evaluated as the following, Cymoxanil determined at wave length detection 240 nm with mobile phase methanol: water (95: 5 v/v) and flow rate 0.5 ml /min with retention time 4.25 min. However, ethylenethiourea determined at wavelength detection 254 nm with mobil phase acetonitrile: Methanol (70: 30 v/v) with flow rate 1.3 ml / min and the retention time was 1.977 min as illustrated by Gamon et al., (1998) and CIPAC (1995). ICP-MS apparatus obtained from thermo fisher company USA to determine of heavy metals lead, cadmium and arsenic as the following forward power 1550w, nebulizer gas 0.8 L/min, injector 2 mm L.D quartz, interface Ni sampler and skimmer, Q cell He gas flow 4.8 ml / min Q cell KED voltage. 2 V and 75 AS dwell time 300 ms.
2.4 Experiment work
2.4.1 Storage stability test at 54 ± 2 °C
The tested fungicides formulation were stored for 14 days at 54 ± 2 °C by the method of (CIPAC 2000). Samples taken before and after storage to determine the active ingredients and relevant impurities for tested fungicide.
2.4.2 Formulation
Standard preparation of tested fungicides and impurities individual standard solutions were prepared in methanol for each fungicide.
2.4.3 Sample Preparation
All fungicides sample before and after storage extracted with methanol and filtrated to analyze by HPLC to determine Cymoxanil and ethylenethiourea percentage.
Determination of metallic copper and heavy metals: The metallic copper percentage was determined before and after storage according to (CIPAC 1967). Cuprrasing was formed by digestion with nitric acid afford with potassium iodide, cupreous iodide, the latter is titrated with sodium thiosulphate. While lead, cadmium and arsenic determined by ICP-Mass apparatus according to (CIPAC 1970).
Determination of Mancozeb by iodometric titration and the active ingredients (a.i) was determined before and after storage by decomposition with acid. Hydrogen sulphide is removed by silver nitrate (in case Mancozeb modified Copper) or by lead acetate (in case Mancozeb alone). Carbondisulphide (CS2) evolved is absorbed methalonic potassium hydroxide and determined iodometric titration according to modified method of (CIPAC 1973).
2.5 Fungal strains used
Pure cultures of Rhizoctonia solani and Fusarium oxysporium supplied from the department of Fungicides, Bactericides and Nematicides, (Central Agricultural Pesticides Laboratory), Agricultural Research Center (A. R. C.).
2.6 Efficacy tested fungicides on pathogenic fungi
Three different fungicides Dicozeb (Mancozeb), Sand flower (Cymoxail + Mancozeb) and Triomax (Cymoxail + Mancozeb + Copper oxychloride) were tested at concentrations of 1000, 500, 400, 300, 200 and 100 ppm before and after storage and evaluated on radial growth of test fungus by Poisoned Food Technique (Mohanty et al., 2012; Moreno-Gavira et al., 2020). Stock solutions of the fungicides were prepared in sterile distilled water and added separately to get the required concentrations thin mixed with 50 ml of sterilized PDA medium and transferred equally into three Petri dishes. The media allowed solidifying. The plates prepared without fungicides served as control. These plates inoculated with 5-mm disc of 7 days old culture of the test fungi. All dishes incubated at 27-±2 °C for 7 days and radial growth of colony measured when the mycelia of control had filled the petri dishes. Each treatment performed in triplicate
The fungal growth inhibition, which calculated due to treatment against control using the following formula: (Satya et al., 2014).
Inhibition of growth (%) = R-r/R *100.
R is the radial growth of fungal mycelia in the control plate.
r is the radial growth of fungal mycelia in the treated plate.
2.7 Statistical analysis
The concentrations inhibition regression lines drawn according to the method of Finney (1971).
2.8 Effect of some fungicides (EC50) before and after storage at 54 ± 2 °C for 14 days on amino acid proline and polyphenol oxidase enzyme produced by Rhizoctonia solani and Fusarium oxysporium
Fungicides (Mancozeb, Cymoxail + Mancozeb and Cymoxail + Mancozeb + Copper oxychloride) at EC50 of each fungus was added to 50 ml sterilized (PD) medium inoculated with 3 discs (5 mm) of any fungus Rhizoctonia solani or Fusarium oxysporium. Three flasks used as replicates. All flasks were incubated at 27 ± 2 °C. When the mycelial growth covered the surface, in the untreated flask. The mycelial matrix excluded by filtration and mycelial mates were homogenized and carried out.
2.9 Determination of polyphenol oxidase activity
Activity of polyphenol oxidase was determined according to the colorimetric procedure adopted by Esterbaner et al., (1977).
2.10 Quantitative assay of the amino acid proline
Amino acid proline determined according to the colorimetric procedure adopted by Bates et al., (1973).
3 Results and discussions
Data in Table 2 shown the degradation rate of the active ingredient of Mancozeb, when Dicozeb was stored at 54 °C for 14 days. The initial value at initial time (zero time) was 79.42 % before storage and decreased to 76.35 % (i.e 3.86 % loss) after 14 days of storage. In addition, in the same Table 1 result showed that the rate of formation ethylenethiourea (Etu) was 0.07 % at one day while, it increased to 0.122% (i.e 74.3% increase) after 14 days from storage and became conformity with (FAO 2010). (Maximum value of Etu 0.5 %). These results are in agreement with those results obtained by (IARC 2001). Ethylenethiourea content in Ethylenebisdithiocarbamate (EBDCs) fungicides formation depends on the pesticides storage conditions and it increased by increasing temperature, a moisture and length of storage period (Lentza-Rizosh, 1990; Camoni et al., 1998; Loper-Fernande et al., 2017). 0 = Initial time before storage.
Storage periods (days)
Mancozeb 80% (WP)
Ethylenethiourea (Etu %) impurities in Mancozeb 80 % (WP)
0
79.42
0.07
14
76.35
0.122
The data in Table 3 shown the degradation rate of two active ingredients Mancozeb and Cymoxanil when Sand flower 72 % WP formulation was stored at 54 °C for 14 days. The initial value at initial time (zero time) was 63.95 % and 7.98 % and decreased to 63.54 % and 7.81 % (i.e 0.64 % loss for Mancozeb and 2.13 % loss for Cymoxanil), respectively after 14 days of storage at 54 °C. According to FAO specifications FAO (2010) and FAO (2006). The tested fungicide Sand flower 72% WP becomes conformity with FAO. 0 = Initial time before storage. % Etu = Ethylenethiourea in content Mancozeb × 100.
Storage periods (days)
Mancozeb 64%
Cymoxanil (a.i) %
Mancozeb (a.i) %
Ethylenethiourea impurities %
0
63.95
0.111
7.98
14
63.54
0.073
7.81
In addition, Results in Table 3 showed that the rate of formation ethylenethiourea was 0.111% at one day and decreased to 0.073 % (i.e. 34.2 % loss) after 14 days of storage at 54 °C ± 2. It found that Etu (impurity) in Sand flower 72 % were bellowing matching the maximum level (0.5 %) defined by (FAO, 2010).
The data in Table 4 shown the degradation rate of three active ingredients (a.i) Mancozeb, metallic copper and Cymoxanil when Triomax 66% WP formulation was stored at 54 °C for 14 days. The initial value at zero time was 11.83, 28.82 and 3.95 % and decreased to 11.63, 28.23 and 3.89 % (i.e 1.69 % loss for Mancozeb, 2.05 % loss for metallic copper and 1.52 % loss for Cymoxanil), respectively after 14 days of storage. According to FAO specification FAO (1989), FAO (2006) and FAO 2010, respectively. The tested fungicide Triomax 66 % WP become conformity with FAO. 0 = initial time before storage. UND undetectable.
Storage periods (days)
Mancozeb 12 %
Metallic copper 29 %
Cymoxanil a.c 4 %
Mancozeb a.c
Ethylene- thiourea %
Copper a.c %
Lead mg/kg
cadmium
Arsenic mg/kg
0
11.83
0.22
28.82
13.77
UND
3.02
3.95
14
11.63
0.227
28.23
13.59
UND
2.96
3.89
Data in Table 4 illustrated the effect of thermal storage at 54 °C on relevant impurities of Ethylenethiourea (Etu) % for Mancozeb, result showed that the rate of formation Etu was 0.22% at zero time and reached 0.227 % after 14 days from storage (i.e 3.18 % increase) and become conformity FAO (2010) (FAO max 0.5 %).
In addition, in Table 4 relevant impurities of lead, cadmium and arsenic mg/kg for copper oxychloride when Triomax 66 % WP was stored for 14 days at 54 °C. Results indicated that there is slightly decrease in heavy metals lead and arsenic content with extending times of storage. The initial value of lead, cadmium and arsenic for one day were 13.77, UND and 3.02 mg/kg and reached to 13.59, UND and 2.96 mg/kg detected of lead, cadmium and arsenic were 0.5 × X mg/kg, 0.1 × X mg/kg and 0.1 × X mg/kg, respectively. Where X is the copper content g/kg, the lead and arsenic as relevant. Impurities became conformity with FAO specification (1989) before and after storage at 54 °C It is similar that Mohamed et al., (2016) found that the amount of relevant impurities cadmium and arsenic of copper fungicide formulation wettable powder slightly affected by storage temperature and became conformity with FAO specification.
Obtained results were agreement with Suzanne Losage (1980) who reported that the formation of Ethylenethiourea by thermal degradation of (EBDCs) in aqueous media greatly reduced the addition of copper salt. In addition, to evaluate of CS2 and the decomposition of Mancozeb decreased when Mancozeb treated with nitrogen group, ester group or ketone group within the molecular structure. For example; 2,6 dimethylmorpholine in tridemorph; N, N-dialkyl aniline in benalaxyl; metalaxyl and oxadixyl or a cyano group, an amino group and an amide group in Cymoxanil which reduced the generation of (Etu) on long storage of the product and reduced the solublized Ethylenethiourea (Etu) during the manufacture or storage of Mancozeb. These process maintained or increased the fungicidal activity of Mancozeb and is more persistent in the field patents Hiroko-kobayashi et al. (1992); Hala et al., 2016; Simonethi et al., 2020.
3.1 Effect of tested fungicides before and after storage at 54 ± 2 °C for 14 days on the mycelial growth of F. Oxysporium in vitro
Data in Table 5 indicated that generally Triomax (Mancozeb + Cymoxanil + Copper oxychloride) was the most effective fungicide against F. oxysporium. It recorded the lowest EC50 value 4.921 ppm, followed by Sand flower (Mancozeb + Cymoxanil) with EC50 value 116.04 whereas the highest EC50 value found with Dicozeb (Mancozeb). It noticed from above results, the fungicidal activity of Mancozeb increased when it mixed with Cymoxanil and Copper oxychloride by 97.4 times while increased to 39.5 time when it mixed with Cymoxanil only. Vadhera et al., (1997) and Dar et al., (2013) Mancozeb and Zineb proved that they are the inhibitor of mycelial growth at all concentrations from 1000 to 2500 ppm while kapadiya et al., (2013) reported that (Cymoxanil + Mancozeb), (Carbendazim + Mancozeb) were most effective against F. solani and F. oxysporium. On the other hand, the effectiveness of tested fungicides deteriorated as result to storage at 54 ± 2 °C for 14 days with EC50 values increased from 4.921 to 116.04, 191.66 to 169.93 and 150.8 to 814.99 in case of Triomax, Sand flower and Dicozeb, respectively. From going results, it concluded that copper oxychloride increased the effectiveness of Mancozeb and decreased the degradation of Mancozeb under storage conditions. (-): disappear BS: before storage AS: after storage. (Dicozeb: active ingredient (mancozeb). Sand flower: active ingredient (mancozeb + cymoxanil). Triomax : active ingredient (mancozeb + cymoxanil + copper oxychloride).
Concentrations (ppm)
Fungicides
Mycelial growth inhibition percentages
Dicozeb
Sand flower
Triomax
BS
AS
BS
AS
BS
AS
1000
100
58
–
80
–
–
500
75
42
100
78
–
89.5
400
65
28
78
76
100
89.4
300
53
22
70
62
89.2
89.0
200
49
18
59
51
88.9
75
100
41
13
53
52
88.3
72
Control
0.0
0.0
0.0
0.0
0.0
0.0
EC50
191.66
814.99
116.04
150.88
4.921
169.93
EC90
1018.1
6393.5
598.22
1713.8
33.865
476.75
Slope
1.767 ± 0.189
1.433 ± 0.19
1.799 ± 0.249
1.214 ± 0.177
0.833 ± 0.374
1.116 ± 0.265
3.2 Effect of tested fungicides before and after storage at 54 ± 2 °C for 14 days on the mycelial growth of R. Solani in vitro
Data in Table 6 indicated that generally Triomax (Mancozeb + Cymoxanil + Copper oxychloride) was the most effective fungicide against Rhizoctonia solani recorded the lowest EC50 value 39.167 ppm, followed by Sand flower (Mancozeb + Cymoxanil) EC50 value 79.527 whereas the highest EC50 value was found with Dicozeb (Mancozeb). From the above results, it noticed that the fungicidal activity of Mancozeb increased when mixed with Cymoxanil and Copper oxychloride by 73.4 times while this increase was 45.9 times when mixed with Cymoxanil only. On the other hand the effectiveness of tested fungicides were deteriorated as a result to storage at 54 ± 2 °C for 14 days the EC50 values increased from 39.167 to 79.527, 147.064 to 58.138 and 603.52 to 745.011 in the case of Triomax, Sand flower and Dicozeb, respectively. From going results, it concluded that Copper oxychloride increased the effectiveness of Mancozeb and decreased its degradation under storage conditions. (-): disappear BS: before storage AS: after storage. Dicozeb: active ingredient (mancozeb). Sand flower: active ingredient (mancozeb + cymoxanil). Triomax: active ingredient (mancozeb + cymoxanil + copper oxychloride).
Fungicides
Concentrations (ppm)
Mycelial growth inhibition percentages
Dicozeb
Sand flower
Triomax
BS
AS
BS
AS
BS
AS
1000
85
54
–
82
–
–
500
68
47
100
76
–
86
400
60
40
73
68
100
78.5
300
55
32
68
56
80
72
200
47
27
60
47
73
65
100
27
15
53
38
67
58.3
Control
0
0
0
0
0
0
EC50
147.064
745.011
79.527
603.52
39.167
58.138
EC90
285,077
39,373
126.953
2195.4
1150.2
236.55
Slope
0.995 ± 0.173
0.744 ± 0.17
1.456 ± 0.256
1.035 ± 0.175
0.873 ± 0.25
2.103 ± 0.415
Effect of tested fungicides (EC50) before and after storage at 54 ± 2 °C for 14 days on amino acid proline and polyphenol oxidase enzyme produced by R. solani and F. oxysporium.
Data in Table 7 showed that before storage a slight increase found in amino acid proline in mycelium of F. oxysporium treated with Dicozeb. In contrast, a slight decrease in the same amino acid found in mycelium of F. oxysporium in the case of Sand flower and Triomax. On the other hand, the mean of proline amino acid increased with all tested fungicide after storage under 54 ± 2 °C for 14 days from 0.38340, 0.34085 and 0.28968 before storage to 0.72550, 0.74423 and 0.99045 after storage in the case of Dicozeb, Sand flower and Triomax, respectively. The point of view, all tested fungicides slightly increased the activity of polyphenol oxidase in F. oxysporium up on control. The storage of these fungicides under 54 ± 2 °C for 14 days increased the activity of polyphenol oxidase from 0.033, 0.029 and 0.025 before storage to 0.076, 0.071 and 0.070 after storage in the case of Dicozeb, Sand flower and Triomax, respectively. Regarding the above results, it concluded that storage decreased the activity of tested fungicides this indication may be due to increase of the mean proline and poly phenol oxides activity in mycelium of fungi. With another point of view, the present of (Cymoxanil + Copper oxychloride) in mixture of Triomax caused a decrease in degradation of its fungicidal activity on other tested fungicides. BS: before storage. AS: after storage.
Fungicides
Fusarium oxysporium
Mean Activity of polyphenol oxidase
Mean of amino acid proline μg/0.1 g mycelium
Dicozeb
(BS)
(AS)0.033
0.076
0.38340
0.72550
Sand flower
(BS)
(AS)0.029
0.071
0.34085
0.74423
Triomax
(BS)
(AS)0.025
0.070
0.28968
0.99045
Control
0.020
0.34293
Data in Table 8 indicated that means of amino acid proline increased slightly in mycelium of R. solani that treated with Triomax compared with control. On the other hand, the mean of proline increased with all tested fungicides that stored under 54 ± 2 °C for 14 days from 0.42542, 0.44360 and 0.58968 before storage to 0.74550, 0.80022 and 0.82045 after storage in the case of Dicozeb, Sand flower and Triomax, respectively. In addition, the mean activity of polyphenol oxides increased in mycelium as resulting of treatment with all tested fungicides up on control. The highest increase found with Dicozeb followed by Triomax and Sand flower. In contrast, the activity of polyphenol oxides increased in mycelium when treated with the same fungicide that stored under 54 ± 2 °C for 14 days from 0.030, 0.031 and 0.037 to 0.066, 0.070 and 0.085 in case of Dicozeb, Sand flower and Triomax, respectively. Regarding the above results, it concluded that degradation of tested fungicides increased the role of these fungicides in increasing the mean activity of polyphenol oxides and mean amino acid proline in mycelium of tested fungi. BS: before storage. AS: after storage.
Fungicides
Rhizoctonia solani
Mean Activity of polyphenol oxidase
Mean amino acid proline μg/0.1 g mycelium
Dicozeb
(BS)
(AS)0.030
0.066
0.42542
0.74550
Sand flower
(BS)
(AS)0.031
0.070
0.44360
0.80022
Triomax
(BS)
(AS)0.037
0.085
0.58968
0.82045
Control
0.032
0.45253
Generally, the commonly available fungicides are effective against plant pathogens. The obtained results indicated that Mancozeb and combination of fungicides (Mancozeb + Cymoxanil), (Mancozeb + Cymoxanil + Copper oxychloride) had high efficiency to control the pathogen. The alternate application of these chemicals reduced the risk of development of resistant in pathogen similar result recorded by El Habbaa et al. (2002).
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
References
- Agrios, G.N., 2005. Plant Pathology, 5th Edition. Elsevier Academic Press, USA, ISBN: 0120445654, 952.
- Diseases of potato in northern areas during. CDRI-PSPDP, PARC, Islamabad: Pakistan; 1995. p. :38.
- Rapid determination of free proline for water stress studies. Plant Soil. 1973;39:205-207.
- [Google Scholar]
- Efficacy of different fungicides against Fusarium solani causing coriander root rot. (Supplement on Plant Pathology). 2014;9(3):1225-1227.
- [Google Scholar]
- Inhibition of phytopathogenic fungi by essential oil, hydrosol, ground material and extract of summer savory (Satureja hortensis L.) growing wild in Turkey. Int. J. Food Microbiol.. 2006;107(3):238-242.
- [Google Scholar]
- CIPAC (1967). Raised 1974 prepared by British cmmitteeof PAC- Charman H Crossely the Murphy Chemical Co. Ltd.
- CIPAC, 1970. Collaborative international pesticides analytical council limited, P 226–228; 236; 238; 637; 1170.
- CIPAC, 1973. Professional method, prepared by dithiocarbamate panel of PAC-UK Chairman Stevenson (Robinson bros 1 td det of heavy metals).
- CIPAC, 1995. Collaborative international pesticides analytical council limited volume F, P. 128, 148, 152, 164, 435.
- CIPAC, 2000. Collaborative international pesticides. analytical council limited reprinted volume J. p. 27, 120, 128, 131.
- Camoni, J.D., muccio, pontecor va A., Hip, D. C., 1998. Survey of ethylenethiourea (Etu) in ethylenebisdithiocarbamate (EBDCs) fungicide. Ecotoxicol Envirou. Saf. 16 (2) 176-179.
- In vitro study of fungicides and biocontrol agents against Fusarium oxysporum f. sp. pini causing root rot of Western Himalayan fir (Abies pindrow) Sci. Res. Essays. 2013;8(30):1407-1412.
- [Google Scholar]
- In vitro evaluation of some fungicides, commercial biocontrol formulations and natural plant extracts on peanut root rot pathogens, Egyptian. J. Agric. Res.. 2002;80(3):1017-1030.
- [Google Scholar]
- A rapid assay for catechol oxidase and laccase using 2-nitro-5-thio benzoic acid. Anal. Biochem.. 1977;1977(77):486-494.
- [Google Scholar]
- FAO specification, 1989. FAO specifications for plant production products, copper oxychloride. Food and Agric. organization of the united nations, Rome, 15-17.
- FAO specification, 2006. FAO specification for plant production products, products, cymoxanil, food and agriculture organizations of the united nations, Rome 12-13.
- FAO specification and evaluations for agricultural pesticide. 2010. Manual on development and use of FAO and WHO specification for pesticides. Second revision of the first edition available only on the internet.
- Probit Analysis Statistical (2nd ed,). Cambridge University; 1971.
- Gamon, M.A., pelegri S.R., peris, L., Dela, J.G., coscolla R., 1998. liquid chromatographic determination of five benzoylurea insecticides in fruit and vegetables. JAOAC Iut., 81 : 1037-1042.
- Synergistic interaction of fungicides in mixture. Am. Phytothl. Soc.. 1996;86(11):1273-1279.
- [Google Scholar]
- Gronogrorac, G., schwack W., 2009. Residue analysis of dithiocarbamate fungicides, Trends in Analytical Chemistry., P40 -50.
- Hala, M. Ibrahim; Naglaa, K. Y. and Waheed, A.M., 2016. Influence of storage thermal on some fungicide compounds and their impurities, J. Biol. Chem. Envirom. Sci. vol. 11(1): 305-307.
- Hiroko-kobayashi; mayumi, N.; shida, Osami-Matano and shinko-Goto, 1992. Effect cystein on the stability of ethylenebisthiourea and ethylenebis(dithiocarbamate) in crops during storage and / or analysis. Journal of Agricultural and food chemistry 40 (1): 76-80.
- IARC, 2001. Ethylenethiourea. In some thyrotropic agents. IARC Monographs on the Evaluation of Carcinogenic Risk of Chemicals to Humans, vol. 79. Lyon, France “International Agency for Research on cancer” pp 659- 701.
- Evaluation of fungicides and herbicides against the root rot of okra. The Bioscan.. 2013;8(2):433-436.
- [Google Scholar]
- Khan, R.A., Iftikhar, S., Rafi, A., Riaz, S., Ahmad, I., 1995. Distribution and incidence of tuber diseases of potato in Swat valley. National Seminar on Research and Development of Potato Production in Pakistan, April 23-25, NARC, PSPDP, PARC, Islamabad.
- Larsson, k.S., Arnander C., Cekanova, E., Jellberg, M.K., 1976. Studies of teratagenic effects of dithiocarbamate maneb, mancozeb and propineb, teratology. 14; 171 – 2 – 184.
- Loper-Fernande, O., Pose-Juan, E., Simal-Gandara, J., 2017. Effect of hydrochemistry variable on half-life of mancozeb and on the hazard index associated to the same of mancozeb and ethylenethiourea. Vol. 154, 253-260.
- ethylenthiourea (Etu) in relation to use of ethylenebisdithiocarbamate (EBDCs) fungicides. Rev. Environ. Contamination Toxicol.. 1990;115:1-39.
- [Google Scholar]
- Storage effect on degradation of copper fungicide and its relevant impurities Egyptian. Sci. J. Pesticide. 2016;2(1):15-22.
- [Google Scholar]
- vitro antifungal efficacy study of plant leaf extracts against three dermatophytes CIB Tech. J. Microbiol.. 2012;1(2–3):27-32.
- [Google Scholar]
- Moreno-Gavira, A.; Dianez, F., Sanchez-Montesinos, B., Santos, M., 2020. Paecilomyces variotii as a plant-growth promoter in horticulture Agronomy 10, 567.
- Fusarium crown and root rot of tomato and control methods. Plant Pathol. J.. 2004;3:9-18.
- [Google Scholar]
- Ong, K., 2011. Basic plant pathology training pathogenic agents. Agi Life Extension Texas, pp 1.
- Peerzada, S.H.; Viswanath, H.S., Bhat, K.A., 2020. In-vitro on the effect of fungicides against mycelial growth and sporangial germination of Phytophthora infestans (Mont) de Bary causing late blight of potato. Int. J. Chem. 8(1): 2069-2075.
- Reinprecht, L., 2010. Fungicides for Wood Protection-World Viewpoint and Evaluation/Testing in Slovakia. pp 95–122.
- First report of Fusarium oxysporum f. sp. lycopersici race 3 on tomato in Brazil. Fitopatologia Brasileira. 2005;30:426-428.
- [Google Scholar]
- Satya, P.R., Manisha S.P., Khushboo S., 2014. Evaluation of the antifungal activity of the potent fraction of hexane extract obtained from the bark of Acacia nilotica. Int. J. Sci. Res.) IJSR) vol 3 issue (10) 730–738.
- Influence of nutrient solution concentration and a root pathogen on tomato root growth and morphology. Sci. Hortic.. 2003;97:109-120.
- [Google Scholar]
- Simonethi, Giulia, Castellani, Federica, Carmela, pomata, Francesca and Sonego, Elisa, 2020. Determination of mancozeb, a pesticide used worldwide in agriculture: comparison among GC, LC, and CE current Analytical chemistry vol 16, No 8 PP. 1041-1053.
- Physicochemical and biological properties metalaxyl, Indain. J. Mycol Pathol.. 1982;12:287-294.
- [Google Scholar]
- Sneh, B., Burpee, L., Ogoshi A., 1991. Identification of Rhizoctonia species. The American Phytopathological Society, Minnesota. Spray Oils against the Sale Parlatona Zizphus 2nd Nat. Conf. of Pests and Dis .of Veg. and Fruits Ismailia, Egypt, 367–376.
- Salicylic acid induced resistance to Fusarium oxysporum f. sp. lycopersici in tomao. Plant Physiol. Biochem. 2009;47:642-649.
- [Google Scholar]
- Effect of cupric ions on the analysis of ethylenebis(dithiocarbamate) residues in tomato juice. J. Assoc. Off. Analy. Chemi.. 1980;63(1):143-144.
- [Google Scholar]
- Non-target effect of fungicides on Rotylenchulus reniformis and Fusarium solani causing root rot of French bean. Adv.. In. Pl. Sci.. 1997;10(1):181-185.
- [Google Scholar]
