COMMENTARY

Insights into differential biomining traits of Indian copper sulphides

Sharadindra Chakrabarti and Ranen Sen

Metal solubilization in microbiological Sulphobacillus sp., Acidithiobacillus fer- nantase and H2SO4 at pH 1.5–1.9 was leaching of copper sulphides is promoted rooxidans, Acidithiobacillus thiooxidans considered. Licananbutase lipoprotein, by synergism of microbe consortium. sp., Acidithiobacillus caldus, Acidithi- an active agent of Licanantase, is obser- Low-grade copper sulphides (chalcopy- obacillus ferridurans and Acidithiobacil- ved in secreted protein fractions of At. rite, pyrite) in India, when subjected to lus ferrivorans, whereas Archaea ferrooxidans and At. thiooxidans. The in- bacterial leaching/biomining in acidic includes Acidianus sp., Ferroplasma sp., itial fractions of 3.5–30 kDa indicate an medium show differential responses. Metallosphaera sp. and Thermoplasma increment in copper recovery. The rea- Such reaction occurs due to geological/ sp. Till today, species of Acidithiobacil- gent contains 5–99% of Licanantase geochemical environments of ore forma- li – At. ferrooxidans, At. thiooxidans, At. lipoprotein along with 1–95% H2SO4 at tion. The co-existence/symbiotic interac- ferriphilus, At. albertensis, At. ferrivo- pH 0.8–3.0. In India, working pH ranges tions of bacterial species along with P–T rans, At. ferridurans, At. sulfuriphilus between 1.4 and 1.9, depending on ore conditions are equally responsible for and At. caldus – are observed. The most specificities. The above reagent has to be their characteristic behaviour. Igneous influential metal bioleaching microbes added at a concentration of 0.01– activities associated with formation of are At. ferrooxidans and At. thiooxidans. 100 mg/l. For ores of four deposits Singbhum (Jharkhand)–Malanjkhand (Mad- Microbial leaching can be direct or in- worked with, the concentration was of hya Pradesh) copper deposits influence direct1. At. ferrooxidans and At. thioox- higher order, varying from 70 to 80 mg/l, the rate of bioleaching of refractory chal- idans are capable of catalysing oxidation whereas Licanantase lipoprotein was copyrite. This rate is higher in sedimen- of reduced sulphur compounds (sulphide, around 87% with 90% H2SO4. tary remobilized deposits of Khetri– elemental sulphur, thionates, etc.), gene- In chalcopyrite–pyrite-dominated low- Rajpura Dariba–(Rajasthan) Ambamata rating sulphuric acid as the final product. grade ores of India, copper recovery (Gujarat). It is found that microbes han- Both these bacteria reduce sulphite and improved by around 9–11%. The Chilean dle leaching problems within genetically thiosulphate as intermediate products experts, however, claimed a progress of defined abilities. that directly/indirectly solubilize metal- up to 20%. World over, 80% copper is produced associated sulphides. This lowers the Molecular genetic studies of Acidithi- through conventional processing (crush- formation of passivating substances on obacilli are time-consuming because of ing, grinding, floatation, fusion-conver- reactive ore surfaces, oxidizing elements long generation time, low yields in liquid sion of concentrates, electrolytic directly by transferring electrons from media and poor growth on solid media. refining). However, this is limited to ore to biomass. At. ferrooxidans can So was earlier accumulated in low-tem- high-medium grade ores, specificities of catalyse oxidation of Fe2+ to Fe3+ that perature bioleaching of chalcopyrite and ore deposits and processing plants. There strongly oxidizes sulphides/other com- associated ores with psychrotolerant pure are none-the-less valuable, relatively pounds whose oxidation is required. and mixed cultures, primarily causing low-grade resources which are sub- Dopson et al.2 expressed doubts about passivation of copper surface. Studies on economic and remain unexploited for the incapability of organomixotroph Fer- microbial ecology of bioleaching till date lack of effective technology. However, roplasma strains to fix carbon dioxide. have relied primarily on standard tech- these ores could be recovered through Leptospirillum, as member of the bio- niques of molecular genetics, especially recent developments in microbial ecology leaching consortium, is providing them in the spatio-temporal distribution of and metabolic processes of biomining/ with fixed carbon3. microorganisms in heap bioleaching. bioleaching, using tools of comparative Bioleaching of Indian ores describes Presently, emerging fields of compara- genomics/metagenomics/bioinformatics. heap bioleaching in acid medium tive genomics and metagenomics have Complete genomic sequences of Acidi- (H2SO4) at pH 1.4–3.5. Incorporating se- broadened our understanding of microbial thiobacillus ferrooxidans and Acidithi- lected biomass with increased oxidizing ecology and metabolic processes of bio- obacillus thiooxidans have led to a better property has been attempted. However, it leaching. understanding of biochemical pathway is difficult to maintain selectivity in Bioinformatics of the genome seque- predictions and physiology of iron– Indian heaps. Increasing heap tempera- nce of At. ferrooxidans has led to several sulphur oxidation for higher copper solu- ture to 50°C to accelerate bioleaching is metabolic regulatory models; some were bilization. impractical for cost constraints. Organic validated in part through experimental Low-grade copper in chalcopyrite of carbon is further added to improve studies like sulphur and iron uptake and Malanjkhand–Khetri–Amba–Mata–Sing- the rate of leaching; but cost nullifies assimilation4,5, carbon metabolism6, etc. bhum may be subjected to bioleaching implementation possibilities. Another Comparisons on a large scale of genomes through direct/indirect microbial actions. impending factor is maintaining carbon addresses fundamental questions, such as Microbes essential for biomining belong under varying heap conditions, because the number of functional genes, identifi- to the bacterial domain of Archaea, that of specificities of ore of a particular cation of species-specific genes, distribu- are acidophilic and chemolithotrophic. region. tion of genes among functional families, In biomining, there are bacteria of For improvement in heap bioleaching etc. Functional abilities of bacteria are Acidiphillium sp., Leptospirillum sp., as executed in Chile, addition of Lica- in fact limited by genomic characters.

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Recent reports7 on Sulfobacillus thermo- lite-granodiorite of early Proterozoic tamination in the copper belt of Ghatsila. sulfidooxidans, At. ferrooxidans and At. age10. The Amba–Mata deposit is a stra- At. ferridurans has similarities with the caldus reveal how the extent of functional ta-bound exhalative sedimentary deposit, gene sequence of At. ferrooxidans; hence abilities depends on genomic attributes. metamorphosed11. Copper mineralization it was earlier incorrectly identified as At. Comparative genomics helps consolidate in Khetri–Kolihan belt is sedimentary– ferrooxidans. gene identification. For example, in iron diagenetic, later remobilized and meta- The whole genome sequencing of oxidation specific to At. ferrooxidans, morphosed12. different Acidithiobacilli species has the potential genetic differences between Various members of the consortium, provided new insights into variegated various Acidithiobacillus species are proactive in leaching, form separate clus- functions. Until now nine At. ferroox- combined to develop evolutionary mod- ters and may or may not be mutually in- idans strain genomes are available15. At. els of the respective genomes and mutual teractive at the initial stage of heap ferrooxidans observed in Indian deposits species interactions (ecophysiology). If bioleaching; but, as the exothermic reac- has not been genetically analysed. Hence leaching problems (which bacteria can- tion proceeds, members of the consor- it is unknown whether it is a variant not handle alone) are visible, the process tium become mutually interactive. strain. At. ferrooxidans IO-2C genome16 needs to be complemented by additional Leaching operations under the Iron has shown significant genetic variants of leaching abilities of bacteria and Arc- Mountains (Northern California, USA) over 75,000 variants. In Indian mines, it haea. The case of Licanantase lipoprote- metagenomic project have, however, is not known which variant and gene ins, discussed earlier, is a pointer. added more comprehensive understand- functions (like iron and sulphur metabol- Bioleaching experimentation at labora- ing of heap bioleaching13. Heap bio- ism, nitrogen fixation, heavy metal resis- tory/pilot/commercial scales in dumps/ leaching proceeds in three stages. It tance, etc.) we are dealing with. The rate heaps have been done over three dec- arises from temperature increase due to of leaching is dependent on gene func- ades. In the late 1980s, the first copper exothermic biological oxidation of (Fe tions. Comparison of genomes addresses dump leaching was attempted at Khetri. II) and S°. An early stage of leaching basic questions such as the number of However, it did not work due to impro- is found at 30–40°C by mesophilic functional genes, identification of spe- per leaching pad design. In the early organisms such as At. ferrooxidans, At. cies-specific genes, distribution of genes 1990s, copper heap/dump leaching- thiooxidans and Sulfurisphaera. As tem- among functional families, mechanisms cementation plant was established indi- perature begins to rise and soars to 40– of reshuffling of genomes, rate of diver- genously at Malanjkhand to utilize sub- 55°C, Leptospirillum, At. caldus and gence of sequence, etc.17. Genomic anal- economic resources, and cement copper Ferroplasma groups are found prevalent. ysis can confirm whether bacteria in produced was fed into the copper conver- At the final stage at 55–65°C or higher, heap can oxidize Fe2+ and sulphur, say at ter at Khetri Flash Smelter. dominantly active Sulpholobus, Alicyc- pH 1.0, and cope with the extreme envi- The thin layer leaching of Peruvian lobacillus and proactive Ferroplasma ronment. Comparative genomic study copper mines was attempted at Ma- (Archaea) flourish. can identify genetic differences between lanjkhand. The ore was cured with strong Consortium of At. ferrooxidans and At. mutually interactive species in leaching acid and agglomerated before being thiooxidans with addition of Licanantase heaps and allow prediction of mutual in- heaped. Sliced heaps gave better results8. protein can remove ~40% copper from teractions between species18. Microbes The ore in heap was cured with 15% well-organized heaps in 45 days. The must be analysed through bioinformatics H2SO4 and wetted for long as feasible, figure is significantly less if a different to unravel the genomic structure. This is before leaching commenced. The higher consortium is used. The data are relevant a prerequisite for successful biomining. the period of wetting, better the result. when copper has igneous associations. Once the genomic structure of proactive However, there was no consortium of With sedimentary metamorphosed ores, microbes is available, capabilities of bacteria integrated by At. ferrooxidans recovery could drop to ~30–35% copper functional genes in leaching operations and At. thiooxidans, and pH was varying in 50 days, with other additional reagents may be understood. The said data pre- between 2 and 3. In the initial period, re- remaining the same. In India, where heap sently are, however, non-existent in sults were quite encouraging but soon leaching is set up for bioleaching of low, India. things faded as the days passed, with a marginal-grade, sub-economic resources In this study, geological factor appears recovery of about 50% in 5 years. of chalcopyrite, the final stage of leach- to be the key component in explaining There is a perceptible relationship bet- ing at 55–65°C is not achievable. There- the rate of solubilization of metal in bio- ween geological origin of ore deposits fore, recovery of metal is always in the leaching operations. The key component and rate of bioleaching. The latter is re- lower range. The absence of third/final is the presence or absence of igneous lated to geology, genesis of deposits and stage of bioleaching is characteristic of activity during ore formation. mode of evolution of the respective Indian copper biomining. Low mineral genomes of mutually interactive micro- surface availability to bacterial mass, bial species in the consortium. primarily on account of pervasive rock 1. Rawlings, D. E., Microb. Cell Fact., Fluid inclusion studies on Singbhum matrix, is probably another cause of less 2005, 4, 13. 2. Dopson, M., Baker-Austin, C. and Bond, copper ore9 show that copper mineraliza- metal recovery. 14 P. L., Microbiology, 2005, 151, 4127– tion is of magmatic/metasomatic paren- At. ferridurans is located only in 4137. tage, or evolved meteoric water at low Singbhum because of the influence of 3. Holmes, D. and Bonnefoy, V., In Bio- water/rock ratio after its interaction with uranium around Jaduguda. At. caldus is mining (eds Rawlings, D. and Johnson, granitic pluton. The Malanjkhand copper more prevalent than At. ferridurans. The B.), Springer-Verlag, Berlin, Germany, deposit occurs within calc-alkaline tona- latter probably indicates radioactive con- 2007, pp. 281–307.

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4. Valdes, J., Veloso, F., Jedicki, E. and 11. Patwardhan, A. M. and Oka, S. S., In 18. Valdes, J., Pedroso, I., Quatrini, R. and Holmes, D., BMC Genom., 2003, 4, Syngenesis and Epigenesis in the Forma- D. S., Hydrometallurgy, 2008, 94, 180– e51. tion of Mineral Deposits (eds Wausch- 184.

5. Quatrini, R., Jedlicki, E. and Holmes, D. kuhn, A., Kluth, C. and Zimmermann, R. ACKNOWLEDGEMENTS. We thank Dr S., J. Ind. Microbiol. Biotechnol., 2005, A.), Springer, 1984, pp. 102–119. Ricardo Badilla (formerly CIMM, Santiago, 32, 606–614. 12. Sarkar, S. C. and Dasgupta, S., Mineral. Chile) for technical advice and Dr Ranjit 6. App-Ayme, C. et al., Hydrometallurgy, Deposita, 1980, 15, 117–137. Choudhuri (formerly RSMM, Udaipur) for his 2006, 83, 273–280. 13. Demargasso, C. S. et al., Hydrometallur- advice on the working out of the main theme 7. Panyushkina, A. E. et al., Sci. Rep., gy, 2005, 80, 241–253. during drafting.

2019, 9, 15069. 14. Miyauchi, T., Kouzuma, A., Abe, T. and 8. Mukhopadhyay, A. D. and Sen, R., De- Watanabe, K., Microbiol. Resour. An- Sharadindra Chakrabarti* is in the De- velopments in biotechnology: microbial nounce., 2018, 7, eo1028–18. partment of Geology, Sister Nibedita leaching and its environment, VU, WB, 15. Zhang, Y., Zhang, S., Zhao, D., Ni, Y., Government College, Hastings House, India, 2003, p. 279. Wang, W. and Yan, L., Microorganisms, Alipore, Kolkata 700 027, India and 9. Mishra, B., Pal, N. and Ghosh, S., J. 2020, 8, 2. Geol. Soc. India, 2003, 61(1), 51–60. 16. Fariq, A., Blazier, J. C., Yasmin, A., Ranen Sen, formerly at Department of 10. Sarkar, S. C., Kabiraj, S., Bhattacharyya, Gentry, T. J. and Deng, Y., 2019, 9, Metallurgical Engineering, Colorado S. and Pal, A. B., Mineral. Deposita, 13049; www.nature.com/scientificreports School of Mines, USA. 1996, 31, 419–431. 17. Hardison, R. C., PLoS Biol., 2003, 1, e58. *e-mail: [email protected]

The re-emerging Karnal bunt disease of wheat and preparedness of the global wheat sector

S. K. Bishnoi, Sudheer Kumar and G. P. Singh

Karnal bunt caused by smut fungus Tille- India, including the range of grain infec- due to Karnal bunt has been quantified to tia indica (syn. Neovossia indica (Mitra) tion for each state. The percentage of range between 0.01% and 1% in India Mundkur), is one of the major fungal infected samples is high; this should be a and Mexico4,7,8. Owing to this, in the be- diseases of wheat that is considered to matter of concern for the Indian wheat ginning, the disease was assumed to be have a high potential of re-emergence, sector and calls for stricter internal qua- of intermediate economic significance particularly in the North Western Plains rantine. only9. The indirect losses caused by Kar- Zone (NWPZ) of India1. It is named after The rise in infection percentage in re- nal bunt also pertain to the rejection of the Karnal district of Haryana, where it cent years has been primarily attributed wheat lots containing more than 3% of was first discovered in 1931 by Mano- to the adaptability of the pathogen to the infected grains for human use10. There- ranjan Mitra2. Although in the NWPZ prevailing weather fluctuations and ab- fore, it is a peculiar wheat fungal disease Karnal bunt has been historically present sence of immunity in wheat mega- that, unlike others (rusts, mildews, etc.), in non-epidemic proportions, precipita- varieties. The extensive use of urea causes monetary losses to the growers tion or high humidity at the flowering (nitrogen fertilizers) for higher yield and more by affecting the quality and less by stage (February–March) may increase the more irrigation events are also responsi- reduction in the quantity of the produce. infection percentage as high as up to ble for an increase in the intensity and Nonetheless, the studies of Murray and 40%. During the post-harvest surveys incidence of Karnal bunt disease out- Brennan11 and Stansbury and McKirdy12 conducted under the All India Coordi- breaks. Karnal bunt is also reported to have shown that we should not be nated Research Project on Wheat and have a high propensity of becoming en- misled by the quantum of direct yield Barley from the grain markets in 2019, demic in new geographies across Europe losses, as the former reported that an 32.02% samples from a total of 7321 and Australia, which are at present free economic loss to the tune of AUD were found infected by T. indica. Maxi- from this disease4,5. The associated risk 490,900,000 per annum (17% of wheat mum infected samples were reported becomes exceedingly important as T. in- economy) will accrue to the wheat com- from Haryana (56.69%), followed by dica is a fungus of high quarantine im- merce if T. indica gets an entry into Aus- Jammu and Punjab with 54.85% and portance across the world with more than tralia. Likewise, the latter study expected 45.18% infected samples respectively. 70 countries having quarantine regula- an even higher economic loss that could Fortunately, none of the samples col- tions imposed against it6. Therefore, the increase up to a significant 25%. The lected from Madhya Pradesh, Gujarat, economic losses caused by the fungus losses estimated included those due to Maharashtra and Karnataka was found to should be understood in the form of a reduction in yield, quality and the ex- carry the Karnal bunt disease3. Table 1 non-tariff barrier to the global wheat pected quarantine regulations that will be presents details of post-harvest sample trade, rather than the direct yield losses in place in case of such a scenario. The survey for Karnal bunt for ten states in which are minor only. The loss of yield expected percentage of losses are enough

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