Open access peer-reviewed chapter

Characterisation of Endo-Polygalacturonases Activities of Rice (Oryza sativa) Fungal Pathogens in Nigeria, West Africa

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Adekunle Odunayo Adejuwon, Marina Donova, Victoria Anatolyivna Tsygankova and Olubunmi Obayemi

Submitted: September 2nd, 2020 Reviewed: October 27th, 2020 Published: June 30th, 2021

DOI: 10.5772/intechopen.94763

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Abstract

Rice (Oryza sativa) is cultivated in swampy geographical locations of tropical Nigeria, West Africa. Here it is infected by a host of fungal pathogens on the field or contaminated at postharvest. This has led to its loss and reduction in its production in both the national and global market. Lasiodiplodia theobromae and Rhizoctonia solani have recently been identified as the major fungal phytopathogens causing the deterioration of this grain on the field and at postharvest and affecting its production in Nigeria leading to gross capital loss. Hence the need to determine physiological control measures for the eradication of both phytopathogens on the field and at postharvest. In this study, tropical strains of Lasiodiplodia theobromae and Rhizoctonia solani obtained from deteriorated rice (Oryza sativa) were grown in a growth nutrient medium composed of MgSo4.7H20, K2HPO4, FeSO4.7H20, potassium nitrate and pectin at 30°C. Endo-Polygalacturonase activities were produced by the fungal isolates in the growth medium within ten days. The endo-polygalacturonases from both fungi were purified by a combination of ammonium sulphate precipitation, dialysis, gel filtration (on Sephadex G-100 column) and ion-exchange chromatography (on CM-Sephadex C-50 and CM-Sephadex C-25 columns). The molecular weight of endo-polygalacturonase from the Lasiodiplodia theobromae using Sephadex G-100 was estimated as 124,000 Daltons while that of the Rhizoctonia solani was estimated as 92,000 Daltons. The purified endo-polygalcuronase from the Lasiodiplodia theobromae exhibited optimum activity at 30°C and at pH 4.5 while that from the Rhizoctonia solani exhibited optimum activity at 32°C and at pH 5.0. The purified endo-polygalacturonases from both fungi exhibited optimum activities at 0.2% pectin concentration. They were stimulated by Ca2+ but inhibited by ethlylenediamine tetracetic acid (EDTA) and 2,4-dinitrophenol. The purified endo-polygalacturonase from the Lasiodiplodia theobromae lost 80% of its activity within 20 minutes of heat at 80°C. While the purified endo-polygalacturonase from the Rhizoctonia solani lost 82% of its activity within 20 minutes of heat at 80°C. Potassium nitrate as nitrogen source in the defined growth medium with pectin as carbon source supported highest activity of endo-polygalacturonase by the Lasiodiplodia theobromae while ammonium chloride as nitrogen source in the defined growth medium with pectin as carbon source supported highest activity of endo-polygalacturonase by the Rhizoctonia solani. In conclusion, the conditions inhibiting endo-polygalacturonases from Lasiodiplodia theobromae and Rhizoctonia solani capable of degrading the pectin portion of rice (Oryza sativa) can be adapted as feasible control measures limiting the infection and contamination of rice (Oryza sativa) by these phytopathogens on the field and at postharvest. Temperature and pH extreme from 30°C and pH 4.5 will be feasible inhibitory control measures for the growth of Lasiodiplodia theobromae on rice (Oryza sativa) in Nigeria while temperature and pH extreme from 32°C and pH 5.0 will inhibit growth of Rhizoctonia solani on the grain. These physiological conditions will preserve pectin in rice (Oryza sativa) from degradation by these two fungal phytopathogens.

Keywords

  • rice (Oryza sativa)
  • phytopathogen
  • fungi
  • microbial enzymes
  • polygalacturonase
  • endo-polygalacturonase
  • purification
  • characterisation

1. Introduction

Oryza sativa(Asian rice) and Oryza glaberrima(African rice) are species of rice grown all over the world [1]. Rice is the species of the seed of grass. It is a cereal grain consumed mostly in Asia and Africa [2]. It is an agricultural grain which has the third-highest worldwide production only after sugarcane and maize [3, 4] and the world’s most consumed staple food [4]. It is rich in starch, protein, minerals and vitamins but low in calories and fats [5]. Rice is grown in all the geographical zones of Nigeria, West Africa depending on the variety [6]. The area of land used for rice cultivation in Nigeria, West Africa is about 2 million hectares. Nigeria however has the potentials of cultivating about 5 million hectares [7, 8].

Rice is affected by a host of fungal pathogens which include: Magnaporthe griseawhich causes Rice blast; Rhizoctonia solaniwhich causes Rice sheath blight; Cochliobolus miyabeanus(an Ascomycete) which causes brown spot disease in rice [1]. In Nigeria, the major fungal pathogens of rice include: Fusarium moniliformeSheldon which causes Bakanae foot rot disease; Cercospora oryzaeMiyake which causes Narrow brown leaf spot; Rhynchosporium oryzaeHashioka and Rhynchosporium oryzaeYokogi which cause Leaf scald; Rhizoctonia solaniKühn which causes Basal sheath rot; Pyricularia oryzaeCav. which causes Rice blast; Cochliobolus miyabeanusIto Dreschler ex Dastur and Cochliobolus miyabeanusKuribayashi Dreschler ex Dastur which cause Rice brown spot; Lasiodiplodia theobromaewhich causes Root rot disease complex in rice [9, 10].

Endo-Polygalacturonase (EC: 3.2.1.15) also known as Pectin depolymerase, PG, Pectolase, Pectin hydrolase, and Poly-alpha-1,4-galacturonide glycanohydrolase, is an enzyme that hydrolyzes the alpha-1,4 glycosidic bonds between galacturonic acid residues. It degrades pectin by hydrolyzing the O-glycosyl bonds yielding alpha-1,4-polygalacturonic residues [11, 12, 13, 14]. This enzyme has multiple parallel beta sheets which form a helical shape that is called a beta helix. This highly stable structure has numerous hydrogen bonds and disulfide bonds between strands common to all pectin degrading enzymes. The interior of the beta helix is hydrophobic [14, 15]. Exo-Polygalacturonases and Endo-Polygalacturonases have differing hydrolytic modes of action. Endo-Polygalacturonases hydrolyze pectin in a random fashion along the polygalacturonan chain resulting in oligogalacturonides. Exo-Polygalacturonases hydrolyze pectin at the non-reducing end of the polymer resulting in monosaccharide galacturonic acid [14]. Fungal Polygalacturonases are affected by a variety of factors which include: pH, substrate concentration, substrate specificity, and temperature [13]. Phytopathogenic fungi expose plant cell walls to Cell Wall Degrading Enzymes (CWDEs) such as Polygalacturonases [14]. Siddiqui [16] purified a monomeric polygalacturase with molecular weight of 32 kDa and optimum activity at 55°C and at pH 5.0 using Sephadex G-200 and Sephacryl S-100 from thermophilic Rhizomucor pusillusisolated from decomposting orange peels. Anand et al. [17] purified an endo-polygalacturonase using acetone precipitation and gel filtration from a strain of Aspergillusfumigatus. They determined the molecular weight to be 43.0 kDa, stable at a pH range of 7–10 and with optimum activity at 30°C. The polygalacturonase was stimulated by Cu2+ and K+ but inhibited by Ag+, Hg2+ and Ca2+. Doughari and Onyebarachi [18] produced polygalacturonase from Aspergillus flavusisolated from orange peel with maximum activity in the presence of polygalacturonic acid at 35°C and pH 4.5. Carrasco et al. [19] reported from their studies the expression of a polygalaturonase in Pichia pastoriswith an optimum activity 15°C higher than its mesophilic counterpart. According to Thakur [20], Mucor circinelloideswas able to produce an extracellular polygalacturonase in a growth medium with pectin methyl ester (1% w/v) as carbon source and a combination of casein hydrolysate (0.1% w/v) and yeast extract (0.1% w/v) as nitrogen source. Optimum polygalcturonase activity was obtained at pH 4.0.

The aim of this study was to determine the physiological conditions that will inhibit the growth of major and specific fungal phytopathogens of rice (Oryza sativa) in Nigeria, West Africa. The present study will establish the contributions of endo-polygalacyuronases produced by Lasiodiplodia theobromaeand Rhizoctonia solanito the deterioration of rice (Oryza sativa) cultivated in Nigeria. Control measures which include conditions inhibiting endo-polygalacturonases from these specific fungal phytopathogens can be adapted in the cultivation (pre-harvest) and storage (post-harvest) of rice (Oryza sativa) globally.

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2. Materials and methods

2.1 Source and identification of isolates

The tropical fungal strains of Lasiodiplodia theobromaeand Rhizoctonia solanifor this investigation were isolated from deteriorated rice (Oryza sativa) obtained from Cocoa Research Institute, Ibadan, Nigeria. The isolates were identified at the International Institute of Tropical Agriculture, Ibadan, Nigeria. The isolates were cultured on potato dextrose agar on plates and slants.

2.2 Inocula and their culture conditions

Lasiodiplodia theobromaeand Rhizoctonia solaniisolated from deteriorated rice (Oryza sativa) were grown in a fungal growth medium with defined specific nitrogen and carbon sources for growth. The isolates were cultured on plates and slants containing potato dextrose agar at 30°C. Ninety six-hour-old cultures of isolates were used in the investigation [21]. Isolates were cultured in a growth medium composed of MgSO4.7H20 (0.1 g), K2HPO4 (2 g), KH2PO4 (0.5 g), FeSO4.7H2O (1 mg), KNO3 (9.9 g) and pectin (10 g) source (Sigma-Aldrich, USA) per 1 litre of distilled water (The pH of the medium was adjusted to pH 5.0 using 0.1 N HCl and 0.1 N NaOH). Conical flasks (250 ml) containing 100 ml growth medium were inoculated with 1 ml of an aqueous spore suspension containing approximately 6 x 105 spores per ml of each isolate. These were the experimental flasks. Control flasks contained un-inoculated medium. Experimental and control flasks were incubated without shaking at 30°C. Protein content was determined [22].

2.3 Enzyme extraction

Contents of flasks were carefully filtered through glass fibre filter paper (Whatman GF/A) on the tenth day of inoculation of growth medium. Protein content of the filtrates was determined [22]. The filtrates were also assayed for polygalacturonase activity [23, 24].

2.4 Precipitation using ammonium sulphate

The crude enzymes of the isolates was treated with ammonium sulphate (analytical grade, Sigma) within 40–90% saturation. Precipitation was at 4°C for 24 hours. Centrifugation was done at 4000 rpm for 30 minutes at 4°C using a high speed cold centrifuge. The precipitate was re-constituted in 0.2 M citrate phosphate buffer (pH 5.0). Protein content of the precipitated enzyme was determined [22]. Polygalacturonase activity was also determined [23, 24].

2.5 Dialysis

The ammonium sulphate precipitated enzyme of each isolate was dialyzed using acetylated dialysis tubings (Visking dialysis tubings, Sigma) [25] and a multiple dialyser (Pope Scientific Inc. Model 220, USA). The enzyme preparation was dialyzed using 0.2 M citrate phosphate buffer (pH 5.0) at 4°C for 24 hours under several changes of the buffer. The protein content of the dialyzed enzyme was afterwards determined using the Lowry et al.[22] method. Polygalacturonase activity of the dialyzed enzyme was also determined [23, 24].

2.6 Enzyme assay

2.6.1 Endo-polygalacturonase (PG)

Reaction mixture consisted of 1 ml of 0.1% (w/v) pectin in 0.2 M citrate phosphate buffer (pH 5.0) as substrate and 0.5 ml enzyme. Controls consisted of only 1 ml of substrate with no enzyme added. The contents of both experimental and control tubes were incubated at 35°C for 1 hr. The reaction in each tube was terminated with 3 ml of dinitrosalicylic acid reagent (Appendix 1). Thereafter, 0.5 ml of enzyme was added to the controls. The contents of tubes were then boiled for 15 minutes. Optical density readings were taken at 540 nm using a colorimeter. The total reducing sugars released in the reaction mixtures was determined by the Dinitrosalicylate (DNSA) method [23, 24]. One unit of endo-polygalacturonase activity was defined as the amount enzyme in 1 ml of reaction mixture which released reducing sugars equivalent to 100 μg galacturonic acid per minute under specified assay conditions. Specific activity was expressed as enzyme units per mg protein.

2.7 Determination of protein concentration

Using the Lowry et al. [22] method, protein concentration was determined at every stage of the investigation. Reaction was with copper in the presence of alkali (Appendix 1).

2.8 Fractionation of enzyme using gel filtration and ion-exchange chromatography

Endo-Polygalacturonase from Lasiodiplodia theobromaeand Rhizoctonia solaniwere subjected to further purification by gel filtration using a Sephadex G-100 column and ion-exchange chromatogeaphy using CM-Sephadex C-25 and CM-Sephadex C-50 columns. The Sephadex G-100 column was calibrated with proteins of known molecular weight [26, 27].

2.9 Characterisation of the purified endo-polygalacturonases

The effects of temperature, pH, substrate concentrations, certain cations and specific inhibitors on the activity of the purified endo-polygalacturonases from Lasiodiplodia theobromaeand Rhizoctonia solaniwere investigated.

2.10 Effect of temperature

The substrate used was 0.1% (w/v) pectin (Sigma-Aldrich, USA) in 0.2 M citrate phosphate buffer (pH 5.0). The reaction mixture consisted 1 ml of substrate and 0.5 ml of enzyme. Incubation was at a range of 5-70°C for 1 hr.

2.11 Heat stability test at 80°C

The effect of heat (80°C) on the stability of the purified endo-polygalacturonase at different periods, 5, 10, 15, 20 and 25 minutes was carried out. The activity of the heated endo-polygalacturonase was determined by incubating 0.5 ml of enzyme plus 1 ml of 0.1% (w/v) pectin (Sigma-Aldrich, USA) in citrate phosphate buffer (pH 5.0) substrate at 35°C for 1 hr.

2.12 Effect of pH

The substrate used was 0.1% (w/v) pectin (Sigma-Aldrich, USA) in 0.2 M citrate phosphate buffer at different pH values ranging from pH 4.0–8.5. The reaction mixture consisted 1 ml of substrate and 0.5 ml of enzyme. Incubation was performed at 35°C for 1 hr.

2.13 Effect of concentrations of substrate (pectin)

Concentrations which were 0.05–0.3% (w/v) pectin (Sigma-Aldrich, USA) constituted in 0.2 M citrate phosphate buffer (pH 5.0) were used as substrate in this investigation. The reaction mixture consisted 1 ml of substrate and 0.5 ml of enzyme, incubated at 35°C for 1 hr.

2.14 Effects of cations

The cations NaCl and CaCl2 were used at different concentrations (5, 10, 15, 20 and 30 mM) for the activity of the purified endo-polygalacturonase. Each cation was constituted in 0.1% pectin in citrate phosphate buffer (pH 5.0. The reaction mixture was 1 ml of substrate and 0.5 ml of enzyme incubated at 35°C for 1 hr.

2.15 Effects of certain inhibitors

2,4-Dinitrophenol and ethylenediamine tetraacetic acid were the inhibitors used in this investigation. They were prepared at concentrations of 2, 4, 6, 8 and 10 mM in 0.1% pectin (Sigma-Aldrich, USA) in citrate phosphate buffer at pH 5.0. These were the substrates.

2.16 Effects of specific nitrogenous compounds

The nitrogenous compounds used were potassium nitrate, ammonium sulphate and ammonium chloride. Pectin was the constant carbon source in the growth medium. Endo-polygalacturonase activity expressed by the fungal strains of Lasiodiplodia theobromaeand Rhizoctonia solaniwere investigated. Endo-Polygalacturonase activity expressed on the tenth day of incubation at 30°C was recorded.

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3. Results

When strains of Lasiodiplodia theobromaeand Rhizoctonia solaniisolated from deteriorated rice (Oryza sativa) were grown in a defined growth medium containing potassium nitrate as nitrogen source and pectin as carbon source, they expressed endo-polygalacturonase activities at 30°C within ten days. The enzymes were isolated and purified by ammonium sulphate precipitation, gel filtration and ion-exchange chromatography. The purification steps are presented on Tables 1 and 2. Purification of the endo-polygalacturonase from the Lasiodiplodia theobromaeby gel filtration using Sephadex G-100 gave four peaks of absorption designated A, B, C. Only the fractions with peak B expressed endo-polygalacturonase activity. The molecular weight estimate of endo-polyglacturonase produced by the Lasiodiplodia theobromaewas 124,000 Daltons. Purification of the endo-polygalacturonase from Rhizoctonia solaniby gel filtration (Sephadex G-100) gave two peaks of absorption designated D and E. Only the fractions of peak E expressed endo-polygalacturonase activity. The molecular weight estimate of endo-polygalacturonase from Rhizoctonia solaniwas 92,000 Daltons.

FractionTotal Activity (Units)Total Protein (mg)Specific Activity (Units/mg protein)Yield (%)Purification fold
Crude extract412060.268.41001
90% (NH4)2SO4 Precipitation322840.380.078.31.16
Sephadex G-100
Gel filtration
Chromatography
Peak A213910.6201.7951.92.95
Peak B10238.3123.2524.81.80
Peak C9954.6216.324.13.16
CM-Sephadex C-50
Ion-Exchange
Chromatography
Peak Ba9662.834523.45.04

Table 1.

Purification of endo-polygalacturonase from Lasiodiplodia theobromaeisolated from deteriorated rice (Oryza sativa).

FractionTotal Activity (Units)Total Protein (mg)Specific Activity (Units/mg protein)Yield (%)Purification fold
Crude extract161532.250.11001
90% (NH4)2SO4 Precipitation132822.160.182.21.2
Sephadex G-100
Gel filtration
Chromatography
Peak D11213.1361.669.47.2
Peak E9267.4125.157.32.5
CM-Sephadex C-25
Ion-Exchange
Chromatography
Peak Eb8956.5137.755.42.7

Table 2.

Purification of endo-polygalacturonase from Rhizoctonia solaniisolated from deteriorated rice (Oryza sativa).

The purified endo-polygalcuronase from the Lasiodiplodia theobromae(ion-exchange chromatographic fraction - CM-Sephadex C-50) exhibited optimum activity at 30°C and at pH 4.5 while the purified endo-polygalacturonase from the Rhizoctonia solani(ion-exchange chromatographic fractions - CM-Sephadex C-25) exhibited optimum activity at 32°C and at pH 5.0.

The purified endo-polygalacturonases from both the Lasiodiplodia theobromaeand the Rhizoctonia solaniexhibited optimum activities at 0.2% pectin concentration.

The purified endo-polygalacturonases from both fungi were stimulated by Ca2+ They were inhibited by ethlylenediamine tetracetic acid (EDTA) and 2,4-dinitrophenol.

The purified endo-polygalacturonase from the Lasiodiplodia theobromaelost 80% of its activity within 20 minutes of heat at 80°C. While the purified endo-polygalacturonase from the Rhizoctonia solanilost 82% of its activity within 20 minutes of heat at 80°C.

Potassium nitrate as nitrogen source in the defined growth medium with pectin as carbon source supported highest activity of endo-polygalacturonase by the Lasiodiplodia theobromaehowever, ammonium chloride as nitrogen source in the defined growth medium with pectin as carbon source supported highest activity of endo-polygalacturonase by the Rhizoctonia solani.

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4. Discussion

Pectin is found in the tissues of rice (Oryza sativa) [28]. In this investigation, we observed that the fungal pathogens Lasiodiplodia theobromaeand Rhizoctonia solaniisolated from deteriorated rice (Oryza sativa) produced endo-polygalacturonases in a growth medium containing pectin as carbon source and potassium nitrate as nitrogen source of fungal growth at 30°C. In Nigeria, West Africa, Lasiodiplodia theobromaeand Rhizoctonia solaniare fungal pathogens of rice (Oryza sativa) in the field [9, 10, 29]. From the results of this study, we can therefore establish that Lasiodiplodia theobromaeand Rhizoctonia solaniare capable of causing the deterioration of rice (Oryza sativa) at 30°C by breaking down the grain’s pectin portion.

In this research, we observed that the molecular weight of endo-polygalacturonase from our strain of Lasiodiplodia theobromaeusing Sephadex G-100 has an estimate of 124,000 Daltons while that of our strain of Rhizoctonia solanihas an estimate of 92,000 Daltons. Adejuwon et al.[11] reported from their studies a polygalacturonase from Penicillium funiculosumThom. isolated from tomato fruits with molecular weight estimate of 89,100 Daltons. Ajayi et al. [12] in their own investigation purified polygalacturonase from Rhizopus arrhizusFisher isolated from tomato fruits with molecular weight estimates of approximately 166,000 Daltons and 60,260 Daltons. Olutiola [13] reported from his investigation, an extracellular polyglacturonase complex from Penicillium citrinumassociated with internal mouldiness of cocoa (Theobroma cacao) beans having molecular weights of 30,000 and 56,000.

The purified endo-polygalcuronase from our Lasiodiplodia theobromaeexhibited optimum activity at 30°C and at pH 4.5 while that from our Rhizoctonia solaniexhibited optimum activity at 32°C and at pH 5.0. Olutiola [30] reported that Penicillium sclerotigenumYamamoto isolated from rotten yam tuber produced a polygalacturonase with optimum activity at pH 5.0.

The purified endo-polygalacturonases from both fungi exhibited optimum activities at 0.2% pectin concentration. The purified endo-polygalacturonases from both fungi were stimulated by Ca2+ but inhibited by ethlylenediamine tetracetic acid (EDTA) and 2,4-dinitrophenol. Ajayi et al.[31] purified polygalacturonase from Rhizopus arrhizusFisher with optimum activity at pH 4.5, stimulated by Ca2+ but inhibited by ethlylenediamine tetracetic acid (EDTA) and 2,4-dinitrophenol. The purified endo-polygalacturonase from our Lasiodiplodia theobromaelost 80% of its activity within 20 minutes of heat at 80°C. While the purified endo-polygalacturonase from our Rhizoctonia solanilost 82% of its activity within 20 minutes of heat at 80°C. Potassium nitrate as nitrogen source in the defined growth medium with pectin as carbon source supported highest activity of endo-polygalacturonase by Lasiodiplodia theobromaewhile ammonium chloride as nitrogen source in the defined growth medium with pectin as carbon source supported highest activity of endo-polygalacturonase by Rhizoctonia solani. According to Olutiola [32], pectin as carbon source with L-asparagine as nitrogen source of a defined growth medium supported the growth and sporulation of Fusarium oxysporumSchlecht. In conclusion, the conditions inhibiting endo-polygalacturonases from Lasiodiplodia theobromaeand Rhizoctonia solanicapable of degrading the pectin portion of rice (Oryza sativa) observed in this study can be adapted as feasible control measures limiting the infection and contamination of rice (Oryza sativa) by these phytopathogens on the field and at postharvest. Temperature and pH extreme from 30°C and pH 4.5 will be feasible inhibitory control measures for the growth of Lasiodiplodia theobromaeon rice (Oryza sativa) in Nigeria and particularly the preservation of pectin in rice (Oryza sativa) from degradation by this particular phytopathogen. Temperature and pH extreme from 32°C and pH 5.0 will inhibit growth of Rhizoctonia solanion rice (Oryza sativa) and preserve pectin in this grain from degradation by Rhizoctonia solani.

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Acknowledgments

Authors are grateful to the British Mycological Society (BMS), Britain, United Kingdom; the National Academy of the Sciences (NAS) of Ukraine, Ukraine, East Europe; and the Institute of Bioorganic Chemistry and Petrochemistry (IBOCP), Kyiv, Ukraine, East Europe for research supports.

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Conflict of interest

Authors declare no conflict of interest.

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A.1 DNSA reagents for terminating endo-polygalacturonase activity

Reagents [17]

  1. Sodium hydroxide – 10 g

  2. Potassium sodium tartrate – 200 g

  3. Phenol – 2 g

  4. 3, 5-Dinitrosalicylic acid – 10 g

  5. 5% Sodium sulphite – 10 ml

  6. 0.03% Glucose – 10 ml

  7. Distilled water – 1Litre

References

  1. 1. Wikipedia (2020b). Rice. Wikipedia, 2020. Wikipedia, The Free Encyclopedia.https://en.wikipedia.org/wiki/Rice#:~:text=Rice%20is%20the%20seed%20of,especially%20in%20Asia%20and%20Africa. Retrieved 13th September, 2020
  2. 2. Dardanelli Group (2015). Rice, Dardanelli Group.http://www.dardanell.com/cereal-pulses/chickpeas/rice.htmlRetrieved 13th September, 2020
  3. 3. Awika JM. Major cereal grain production and use around the world. ACS Symposium Series. 2011;1089:1-13
  4. 4. Folnovic, T. (2020). National Rice Month. Agrivi 2020.https://blog.agrivi.com/post/national-rice-monthRetrieved 13th September, 2020
  5. 5. United Prime Publications (2018). Journal of Rice Science. United Prime Publications, 2018.https://www.untdprimepub.com/journal-of-rice-science/Retrieved 13th September, 2020
  6. 6. Longtau SR. Multi-Agency Partnerships in West African Agriculture: A Review and Description of Rice Production Systems in Nigeria. In: Rice Production in Nigeria, ODI; 2003 50pp
  7. 7. FAO (2020). Nigeria at a Glance. FAO in Nigeria. In: Food and Agricultural Organization of the United Nations, 2020.http://www.fao.org/nigeria/fao-in-nigeria/nigeria-at-a-glance/en/Retrieved 13th September, 2020
  8. 8. Oteng, J.W. and Anna, R.S. (1999). Rice Production in Africa: Current Situation and Issues. In: International Rice Commission Newsletter, Volume 48 of the Food and Agricultural Organization of the United Nations, 1999
  9. 9. Awoderu VA. Rice diseases in Nigeria. PANS Pest Articles and News Summaries. 1974;20(4):416-424
  10. 10. Claudius-Cole AO.Lasiodiplodia theobromaein the root rot disease complex of rice. Journal of Rice Research. 2018;6(4):1000197. DOI: 10.4172/2375-4338.1000197
  11. 11. Adejuwon AO, Oni OA, Olutiola PO. Polygalacturonase from tomato fruits infected withPenicillium funiculosumThom. Journal of Plant Sciences. 2006;1(4):383-387
  12. 12. Ajayi AA, Adejuwon AO, Olutiola PO. Partial purification of polygalacturonase from tomato fruits infected byRhizopus arrhizusfisher. Journal of Plant Sciences. 2007b;2(2):216-221
  13. 13. Olutiola PO. Extracellular polygalacturonase enzyme complex fromPenicillium citrinumThom. Associated with internal moldiness of cocoa (Theobroma cacao) beans. Acta Phytopathologica Academiae Scientiarum Hungaricae. 1982a;17(2-4):239-247
  14. 14. Wikipedia (2020a). Polygalacturonase. Wikipedia, 2020. Wikipedia, The Free Encyclopedia.https://en.wikipedia.org/wiki/Polygalacturonase#:~:text=Polygalacturonase%20(EC%203.2.,bonds%20between%20galacturonic%20acid%20residues. Retrieved 13th September, 2020
  15. 15. Proteopedia (2018). Polygalacturonase. Proteopedia, 2018. ISPC, Weizmann Institute of Science, Israelhttp://proteopedia.org/wiki/index.php/PolygalacturonaseRetrieved 13th September, 2020
  16. 16. Siddiqui MA, Pande V, Arif M. Production, purification and characterization of polygalacturonase fromRhizomucor pucillusisolated from decomposting orange peels.Enzyme Researchvolume 2012, article ID 138634. In: 8 pages doi:10.1155/2012/138634. 2012
  17. 17. Anand G, Yadav S, Yadav D. Purification and characterization of polygalacturonase fromAspergillus fumigatusMTCC 2584 and elucidating its application in retting ofCrotalaria junceafiber. 3 Biotechnology. 2016;6:201. DOI: 10.1007/s13205-016-0517-4
  18. 18. Doughari JH, Onyebarachi GC. Production, purification and characterization of polygalacturonase fromAspergillus flavusgrown on orange peel. Applied Microbiology. 2019;4(3). DOI: 10.4172/2471-9315.1000155
  19. 19. Carrasco M, Rozas JM, Alcaino J, Cifuentes V, Baeza M. Pectinase secreted by psychrotolerant fungi: Identification, molecular characterization and heterologous expression of a cold active polygalacturonase fromTetracladium sp. Microbial Cell Factories. 2019;18:45. DOI: 10.1186/s12934-019-1092-2
  20. 20. Thakur A, Pahwa R, Singh S, Gupta R. Production, purification and characterization of polygalacturonase fromMucor circinelloidesITCC 6025.Enzyme ResearchArticle ID. 2010;170549, 7 pages. DOI: 10.4061/2010/170549
  21. 21. Adejuwon, A. and Tsygankova, V. (2020). α-Amylase Production by Toxigenic strains of Aspergillus and Penicillium In:Aflatoxin B1 Occurrence,Detection and Toxicological Effects(Long, X-D. Ed). Intechopen Limited, London, United Kingdom. ISBN: 978-1-83880-256-1; Print ISBN: 978-1-83880-255-4 DOI: doi:10.5772/intechopen.86637
  22. 22. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement with the folin phenol reagent. Journal of Biological Chemistry. 1951;193:265-275
  23. 23. Miller GL. Use of Dinitrosalicylic acid reagent for the determination of reducing sugar. Analytical Chemistry. 1959;31:426-432
  24. 24. Olutiola PO. Cell wall degrading enzymes associated with the deterioration of cocoa beans byPenicillium steckii. International Biodeterioration Bulletin. 1983;19:27-36
  25. 25. Whitaker DR, Hanson KR, Datta PK. Improved procedure for purification and characterization of Myrothecium cellulase. Canadian Journal of Microbiology and Physiology. 1963;41:671-696
  26. 26. Andrews P. Estimation of the molecular weights of proteins by Sephadex gel-filtration. Biochemical Journal. 1964;91(2):222-233
  27. 27. Olutiola PO, Cole OO. Production of a cellulase complex in culture filtrates ofAspergillus tamariiassociated with mouldy cocoa beans in Nigeria. Physiologia Plantarum. 1976;37:313-316
  28. 28. Shibuya N, Nakane R, Yasui A, Tanaka K, Iwasaki T. Comparative studies on cell wall preparations from rice bran, germ. and endosperm.Cereal Chemistry. 1985;62(4):252-258
  29. 29. Reddy OR, Sathyanarayana N. Seed-borne fungi of Rice and quarantine significance. In:Major Fungal Diseases of Rice. Dordrecht: Springer; 2001. DOI: 10.1007/978-94-017-2157-8_24
  30. 30. Olutiola PO. Polygalacturonase and pectin lyase ofPenicillium sclerotigenum. Nigerian Journal of Microbiology. 1982b;2(2):154-167
  31. 31. Ajayi AA, Adejuwon AO, Olutiola PO. Characterization of polygalactturonase from tomato (Lycopersicon esculentummill.) fruits infected byRhizopus arrhizusfisher. IFE Journal of Science. 2007a;9(2):149-154
  32. 32. Olutiola PO. Growth, sporulation and production of pectic and cellulolytic enzymes inFusarium oxysporum. Transactions of the British Mycological Society. 1978;70(1):109-114

Written By

Adekunle Odunayo Adejuwon, Marina Donova, Victoria Anatolyivna Tsygankova and Olubunmi Obayemi

Submitted: September 2nd, 2020 Reviewed: October 27th, 2020 Published: June 30th, 2021