Microbioz India, September 2021 Lab 2021 digital

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Contact: Email : [email protected] Tel: +91-9082974025 Microbioz India, September 2021 Edition | 5

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33 51 22 12 24 Cover Story 12 Pharmaceutical Mycelia: A story of statins Featured Article 18 28 Conducting Research on Highly Pathogenic Viruses Using Virus 4238 Pseudotypes: The Influence of Ultrapure Water on Data Quality Business News IBM Research and Arctoris accelerate closed loop drug discovery with AI and Cloud Product Launches All You Need to Know About the Zebrafish Model in Biomedical and Applied Research Microbioz India, September 2021 Edition | 8

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ycelial cultures were obtained from fragments of F. officinalis fruiting bodies. The taxonomic position of the mushroom mycelium was confirmed using the PCR method. The presence of organic compounds was determined by HPLC–DAD analysis. The cover story of this month titled: “Pharmaceutical Mycelia: A story of statins” contributed by LGC Mikromol. As the story suggests Cholesterol: we all know the word, and its negative associations. So is all cholesterol bad? It’s not that simple – the fatty substance known as cholesterol is not only ingested through food, but also created by the liver, and has crucial functions without which we wouldn’t be alive. These functions include contributing to cell wall structure, forming digestive bile acids in the intestine, and enabling the production of vitamin D and certain hormones. Cholesterol is carried round the body in two types of lipoproteins, low-density lipoprotein (LDL) and high-density lipoprotein (HDL). The edition has a featured article entitled “Conducting Research on Highly Pathogenic Viruses Using Virus Pseudotypes: The Influence of Ultrapure Water on Data Quality” contributed by Sartorius. From 2013 to 2016, more than 11,000 people died of Ebola virus infections in West Africa. This most recent epidemic shows how dangerous so-called emerging viruses can be. Virus pseudotypes can be used to easily investigate the entry pathways of such viruses. Novel viruses, termed emerging viruses, pose a threat to human health of a magnitude that should not be underestimated. Dear subscribers, we could not imagine anything better than to have your essential criticism and proposals since they help us to concoct much better material each time. For any suggestion and feedback please feel free to write: [email protected] Disclaimer: Microbioz India, September 2021 Edition | 10 Neither the Microbioz India nor its publishers nor anyone else involved in creating, producing or delivering the Microbioz India (in printed, web or CD format) or the materials contained therein, assumes any liability or responsibility for the accuracy, completeness, or usefulness of any information provided in the Microbioz India Magazine (in printed, web or CD format), nor shall they be liable for any direct, indirect, incidental, special, consequential or punitive damages arising out of the use of the Microbioz India magazine.

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Cover Story Pharmaceutical Mycelia: A story of statins Cholesterol: we all know the word, and its negative associations. So is all cholesterol bad? It’s not that simple – the fatty substance known as cholesterol is not only ingested through food, but also created by the liver, and has crucial functions without which we wouldn’t be alive T hese functions include contributing to cell HDL absorbs cholesterol and carries it back wall structure, forming digestive bile acids to the liver, which can regulate cholesterol levels in the intestine, and enabling the and clear some from the body if necessary. LDL production of vitamin D and certain hormones. carries the majority of your body’s cholesterol and Cholesterol is carried round the body in two types can cause build-up in blood vessels when there is of lipoproteins, low-density lipoprotein (LDL) and too much of it in your blood – potentially leading high-density lipoprotein (HDL). to heart attack or stroke. Microbioz India, September 2021 Edition | 12

Cover Story One of the most By 2012 atorvastatin/Lipitor had become the top- commonly prescribed selling pharmaceutical of all time , and medications in the world, even in the latest list of most prescribed drugs in statins are a group of the US (2021 list – using 2018 prescription data), it agents used to both lower still sits at the no 1 spot, at 112 million the level of LDL prescriptions. cholesterol in the blood and for Magic mushrooms their antithrombotic and anti- Statins are secondary metabolites – low-weight inflammatory molecular compounds produced by effects. These actions microorganisms and plants – produced from help to prevent blood filamentous fungi. clotting and build-up on blood vessel walls, thus Microbioz India, September 2021 Edition | 13 reducing the risk of heart attacks and strokes. Common statins today include atorvastatin, fluvastatin, simvastatin and rosuvastatin. History While many statins are now produced synthetically, their natural precursors were predominantly produced from Aspergillus and Penicillium fungus species. In the 1970s Japanese scientist Akira Endo began investigating HMG CoA in microorganisms and discovered mevastatin from the fungus Penicillium citrinum. Mevastatin is considered the first statin to be isolated and developed, although it never reached the market. The first statin to market, lovastatin, was isolated from the fermentation broth of Aspergillus terreus in the 1970s by Alfred Alberts, who worked for Merck, and made available to the public in 1987 as Mevacor. But it was atorvastatin, first patented by Warner-Lambert in 1986, that was to take statins stratospheric. A five-year study published in 1994 researching the effects of Merck’s synthetic lovastatin alternative, simvastatin, on patients with high cholesterol and heart disease found a 35% reduction in their cholesterol and 42% lower chance of dying of heart attack in the period. Early results comparing atorvastatin to simvastatin indicated that Warner-Lambert’s drug was both more potent and had fewer side effects, so in 1996 atorvastatin reached market under the name Lipitor, in a joint agreement with Pfizer, who later acquired Warner- Lambert. Lipitor soon became a popular prescription, its arrival coinciding with an increase in average weight and cholesterol levels seen across many developed societies, and by 2003 was the best-selling medication in the US.

Cover Story As many secondary metabolites serve as defence These two compounds can then be modified to give semi-synthetic statins – pravastatin, which molecules against other microbes, despite being is derived from mevastatin, and simvastatin, derived from fungi they often have antimicrobial or which is derived from lovastatin. The third class of statins are fully synthetic, antifungal activity. including fluvastatin, rosuvastatin and ator Fungi are an incredibly diverse group of eukaryotic vastatin, among others. Fluvastatin was the first fully synthetic drug to be developed. Atorvastatin is organisms once considered like plants, though commonly sold as a calcium complex, featuring two molecular evidence has shown that they are atorvastatin ions and a calcium ion. Rosuvastatin is marketed in the same way. actually more closely related to animals Structurally, the natural and semi-synthetic than to plants, with a shared common ancestor compounds are similar, sharing a chiral β-hydroxy- who split from plant life. There are clues to their δ-lactone (or in the case of pravastatin an open chain analogue) and a hexahydronaphthalene distance from plants in their cell walls made of system. Pravastatin has an additional hydroxyl chitin – not found in the plant world but common group on its hexahydronaphthalene group, and this is obtained by microbial hydroxylation of in insects – rather than cellulose, their lack of mevastatin. The addition of this hydroxyl group chlorophyll, production of spores and in the means pravastatin has a much higher solubility in water. The fully synthetic statins possess the same mycelium, the branching filamentous structure of 3,5-cis-dihydroxy acid motif seen in pravastatin, but hyphae. a heterocyclic ring replaces the hexahydronaphthalene structure. Fungi absorb nutrients through this mycelium and Simvastatin possesses an additional methyl group on the butyrate ester, lending the drug a much many exist in a symbiotic relationship with trees, higher bioavailability than lovastatin. To obtain grasses and crops, known as a mycorrhiza. This simvastatin, lovastatin is reduced using lithium hydroxide, resulting in a compound complex association is essential to plant life and the with two hydroxy groups that are susceptible to mycelia of fungi can span entire forests – the largest reaction (Figure 2). Since the hydroxy on the lactone functional group is necessary for enzyme know organism is Armillaria ostoyae, which covers binding, it is protected by organosilicon compound 3.4 square miles (8.8 km2) in the Malheur National TBDMS in the presence of imidazole, leaving the other hydroxy free to react with 2,2- Forest. It is aged between 1900 and 9600 years old. dimethylbutyryl chloride. Some species of fungi are even carnivorous, with mycelia that can digest living organisms such as nematodes: species in the saprotrophic Pleurotus mushroom genus are good examples. While lovastatin was found in A. terreus, it is also produced in some higher fungi, most of which are in the Pleurotus genus. It is most known in P. ostreatus, the oyster mushroom, which contains up to 2.8% lovastatin on a dry weight basis, but can also be found in P. djamor, Imleria badia, and Cantharellus cibarus, among others. Edible mushrooms, meanwhile, have been collected for food for thousands of years, with archaeological evidence that they may have been used as far back as 6800 years ago in Neolithic China. Many mushrooms are psychoactive, which led to roles in medicinal and ceremonial activities in indigenous tribes. Even the common red-and-white spotted mushroom Amanita muscaria or fly agaric, which features in children’s picture books, garden ornaments and video games, is actually hallucinogenic. Chemistry of statins Statins can be divided into three groups based on how they are obtained. As mentioned, mevastatin and lovastatin are naturally occurring statins obtained via fermentation methods. Microbioz India, September 2021 Edition | 14

Cover Story In this case, the lactone group is hydrolysed in the liver to form a hydroxy acid form (Figure 3) which If the hydroxy group isn’t properly protected the bears a considerable similarity to the structure of compound can undergo esterification to give Imp HMG CoA. This similarity in structure allows the B, or be completely removed to give Imp C. statins to fit into the active site of HMG-CoA Simvastatin can also dimerise (Imp D). reductase and competitively inhibit cholesterol synthesis. Mechanism of action Outlook: Mushroom for growth? Statins work by binding competitively to an While statins are already hugely popular drugs, enzyme known as HMG-CoA reductase. This is a rate-controlling enzyme in the mevalonate there is evidence that these fungi-derived pathway, the metabolic process responsible for compounds may have even more medical uses than cholesterol production. The enzyme catalyses the currently confirmed. conversion of metabolic intermediate HMG-CoA Even within their current medical functions, they to mevalonic acid. may have more expansive potential uses: few controlled trials of statins have included patients A clear difference between most of the over 70 years old, because the assumption has been natural statin compounds and the synthetic that their effects would be of limited use in people ones is the presence and absence of a with a shorter life expectancy, and might even be lactone group. This is because mevastatin, damaging. However, a 2020 observational lovastatin and simvastatin are study of 320,000 patients aged 75 years and administered in their prodrug older, free of heart disease and taking statins for the form. Prodrugs are an important first time, found a 25% lower risk for death and a 20% lower risk for cardiovascular death over seven class of drug delivery systems, which provide a means to improve years than those not prescribed statins. A further bioavailability, improve how a drug two studies have also indicated that statin use interacts with cells, and prevent unwanted benefits elderly patients as much, if not more than, side effects. Prodrugs themselves are younger ones. chemically inactive, but after administration Data has also indicated that statin use could help protect against cardiotoxic chemotherapy, with are metabolised into the pharmacologically statins associated with a 55% reduced likelihood of active drug. hospitalisation for heart failure in breast cancer chemotherapy patients over five years in one analysis. Outside cardiovascular use, statins have also been suggested to have therapeutic potential in a wide range of other areas, from dementia to lung cancer. Stroke risk seems to be reduced through statin use, with meta-analysis of 19 studies finding that the prevention of ischemic stroke was striking enough to outweigh any associated risk. Compelling studies also suggest that statins offer ‘endothelial protection…as well as antioxidant, anti-inflammatory and anti- platelet effects’, which could have potential for treatment not only of stroke but also in neurological disorders such as Alzheimer’s, Parkinson’s and MS. Microbioz India, September 2021 Edition | 15

Cover Story This is where Mikromol can help – we support your pharmaceutical quality testing for a wide range of statin APIs and related impurities, with certified reference standards created by our expert chemists. If even some of these many potential additional uses for statins prove to be fruitful, it will further cement their place in the pharmaceutical hall of fame. But one thing’s for sure: these versatile compounds have already changed the world for the better. About the author Lungs are also impacted: statins have also been Dulcie Phipps is an Assistant demonstrated to have potential to lower the Global Product Manager at mortality rate in patients with chronic LGC. obstructive pulmonary disease (COPD), and in A chemist with a passion for population studies long-term statin use has been nature, she grew up in associated with improved overall lung the glens of Perthshire, cancer survival times. Scotland. Phipps’ academic There has also been exploration of prescription of background includes a Marine statins for immune and inflammatory disorders, to Science HNC, an MSc in Chemistry from the University of treat parasitic diseases, and for their antibacterial Glasgow, and a European work placement focusing on properties. Whatever the clinical use, they all have lanthanide chemistry at Uppsala University, Sweden. After one aspect in common: they need high quality, safe completing her degree, she moved to Luckenwalde, and reliable pharmaceuticals to provide patients Germany, to work for LGC. with efficacy and peace of mind. Microbioz India, September 2021 Edition | 16

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Featured Article Conducting Research on Highly Pathogenic Viruses Using Virus Pseudotypes: The Influence of Ultrapure Water on Data Quality This application note was first published by Sartorius- click here for the full application note and references F rom 2013 to 2016, more than 11,000 Furthermore, the surrounding circumstances of a people died of Ebola virus infections in globalised society involving increased travel also West Africa. This most recent epidemic enable dissemination of pathogenic viruses around the world, even before the onset of clinical shows how dangerous so-called emerging viruses symptoms. Within the last 100 years, multiple introductions of emerging viruses into the human can be. Virus pseudotypes can be used to easily population have occurred, which have led to local epidemics or worldwide outbreaks (pandemics; see investigate the entry pathways of such viruses. Table 1). Novel viruses, termed emerging viruses, pose a threat to human health of a magnitude that should not be underestimated. These viruses have acquired the ability to infect In particular, the Ebola virus most recently raised humans either as a consequence of interspecies concerns across the globe, as the West African transmission or of naturally occurring changes in Ebola virus epidemic, which claimed more than the viral genome. Infections caused by emerging 11,000 lives, devastatingly demonstrated the danger viruses often result in serious diseases or even death posed by emerging viruses. Research involving as the human immune system may not be capable emerging viruses is often limited to high- of combatting an unfamiliar virus, especially of containment laboratories of biosafety levels (BSL) zoonotic origin. Various factors promote the 3 and 4 (see Table 1). Working in BSL-4 occurrence and spread of emerging viruses. These laboratories is highly laborious, expensive and include ecological factors, such as deforestation to permitted only at a few locations so that rapid gain new land for development, and also our scientific progress in characterizing viral pathogens lifestyle involving global mobility, as well as and developing antiviral drugs is difficult to achieve international commerce. Thus, the loss of habitat increases the probability that people, pets and farm Given this situation, virus pseudotypes offer an animals will come in contact with wildlife species attractive option for studying the entry of highly originally inhabiting isolated areas and therefore pathogenic viruses into cells safely and efficiently. not previously encountered as natural hosts of emerging viruses. Microbioz India, September 2021 Edition | 18

Featured Article This is possible as not the entire pathogen is The purity level of the laboratory water used analysed, but rather only its components that influences the quality of VSV pseudotypes mediate host cell entry, the envelope proteins. These represent the key to virus entry into cells. After it could be shown in the previous trials that VSV pseudotypes are suitable models for studying In the case of virus pseudotypes, envelope proteins the host cell entry of highly pathogenic viruses, the of highly pathogenic viruses are incorporated into next issue to be clarified was the particular a carrier virus (pseudotyping), which cannot influence that the purity level of the laboratory replicate autonomously, i.e., are replication water used has on the quality of VSV pseudotypes. deficient. Commonly used systems for During production of VSV pseudotypes, various pseudotyping are based on rhabdoviruses (e.g., buffers and solutions are employed that are all vesicular stomatitis virus, VSV; Fig. 1) and prepared with water. However, it must be noted retroviruses. The objective of this study was to that not just any type of water suffices for verify whether the envelope protein-mediated entry preparation of these reagents. Rather, the right of virus pseudotypes reflects host cell entry of choice of purity level must be made for laboratory intact viruses. In addition, this investigation was use. To investigate whether the use of ultrapure designed to determine which influence the level of water yields higher quality VSV pseudotypes, two purity of the reagents used, in this case laboratory batches of VSV pseudotypes were generated in a water, has on the production of virus pseudotypes parallel experiment: One batch was prepared using demineralised water from a central lab water supply It is known that the Middle East respiratory source (conductivity of 3.7 – 4.1 μS/cm at 19°C) as syndrome coronavirus (MERS-CoV, formerly the basic solvent for all solutions and buffers, while called human coronavirus, EMC = hCoV-EMC), a further batch utilising solutions and buffers based binds to the cell surface through the interaction on Arium® Pro VF ultrapure water (conductivity between the viral spike glycoprotein (S) and the 0.055 μS/cm compensated to 25°C) was generated. cellular membrane protein dipeptidyl peptidase 4 (DPP4), thus enabling cell entry [7]. To confirm Microbioz India, September 2021 Edition | 19 whether this also applies in the context of VSV pseudotypes, target cells were transfected with an expression vector for DPP4 or an empty expression plasmid (no receptor). As expected, the directed expression of DPP4 led to a significant increase in the host cell entry of VSV pseudotypes if the pseudotypes had MERS-CoV S embedded in their envelope (see Fig. 2A). Influenza A viruses, the causative agents of influenza disease, require terminal sugar structures, so-called sialic acids occurring as natural modifications on cellular membrane glycoproteins and glycolipids, as receptors to mediate entry into target cells (see literature for further reading [8]). To study whether the incorporation of influenza A viral envelope proteins in VSV pseudotypes also results in sialic acid-dependent cell entry, VSV pseudotypes with H1N1 (1918) HA/NA were employed, and the sialic acids were enzymatically removed from the surfaces of target cells. As expected, removal of sialic acids resulted in a dramatic decrease in the host cell entry of VSV pseudotypes that had H1N1 (1918) HA/NA embedded in their envelope (see Fig. 2B). This finding confirms that virus pseudotypes reflect the entry mechanism of authentic influenza A viruses into cells.

Featured Article The use of pseudotypes also reduces the considerable labour intensity involved in high- EBOV GP and MERS-CoV S as envelope proteins containment laboratories requiring whole-body were examined. Following parallel production of protection suits, as well as the considerable costs VSV pseudotypes by maintaining identical and the limitations entailed by such lab work (e.g., incubation conditions, the target cells were no access to equipment that is not directly located inoculated and the envelope protein-mediated inside the high-containment laboratory). entry of VSV pseudotypes was quantified. This Furthermore, compared with authentic, highly experimental setup demonstrated that host entry pathogenic viruses, virus pseudotypes minimise the (as a parameter for the degree of quality) of the risk of infection of lab personnel following VSV pseudotypes produced using Arium® Pro VF unintentional exposure, thus representing a ultrapure water as the basic solvent for all buffers significant safety aspect. Optimisation of the and solutions was significantly higher compared production process, for example, through the use with that of pseudotypes for which demineralised of highly pure reagents, particularly those based on water was employed in their production. water, can additionally contribute to improving the sensitivity of subsequent test procedures, Conclusion increasing production quantities and thus further lowering production costs In conclusion, it can be stated that virus pseudotypes are important tools for investigating This application note was first published by host cell entry of highly pathogenic viruses. As Sartorius- click here for the full application note these virus pseudotypes do not restrict research on and references. Find out more on the importance such highly pathogenic viruses to BLS-3 or BLS-4 of the quality of ultrapure water, among other laboratories, this enables a larger number of factors, for the production of such virus scientific facilities to conduct such research. As a pseudotypes. result, the cell entry of emerging viruses can be characterised and suitable detection procedures and Other Covid 19 related resources- Sartorius Covid antiviral strategies (medications, vaccines) can be 19 Resource page developed faster. Microbioz India, September 2021 Edition | 20

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Featured Article Sartorius Unveils LIVECell, a Deep-Learning Dataset for Label-Free, Quantitative Cell Segmentation in Nature Methods Source: Sartorius Newsroom  Open-source, manually annotated, expert-validated dataset includes more than 5000 label-free phase- contrast microscopy images acquired by the Incucyte® Live-Cell Analysis System, consisting of more than 1.6 million cells of eight cell types with distinct morphologies  Diversity of the dataset enables its use for more effective training of neural networks for creation of customized machine learning algorithms, leading to more accurate segmentation of additional cell types The Life Science Group Sartorius today While neural networks can learn and adapt to announced publication of an article identify and segment a variety of cells, they first in Nature Methods describing the require training with high quality datasets company’s LIVECell (Label-free In Vitro image representative of the breadth of the cell Examples of Cells) deep-learning dataset for label- morphologies to be encountered. free, quantitative segmentation of live cell images. The open- source dataset includes 5000 label-free phase contrast microscopy images consisting of more than 1.6 million cells of eight cell types with distinct morphologies that have been manually annotated. The set of images includes cells grown from initial seeding densities to fully confluent monolayers, resulting in a large variation in cell size and shape. “The ability to derive IncuCyte-S3-SX5-mAb-Monoclonal- Antibodies-Sartorius physiologically relevant data Achieving accurate segmentation in microscopy from label-free microscopy images is a cornerstone images is essential for quantitative downstream of pharmaceutical research and datasets containing analysis but is a challenging task. Traditional image images of millions of cells facilitate exploration of analysis methods often require tedious algorithm biological phenomena with great statistical power,” customization and rigorous tuning of parameters said Rickard Sjögren, PhD, Senior Scientist, specific to the cell morphology of interest.“The Sartorius Corporate Research. diversity of cell types and confluence conditions “To compensate for a lack of image resolution, captured and annotated in the LIVECell dataset however, sophisticated imaging processing pipelines are necessary to generate the accurate cell- overcomes these challenges by facilitating the by-cell, pixel-by-pixel segmentations necessary to capture subtle changes in cell size, shape and training of deep learning-based segmentation texture, particularly if the goal is to investigate events at the level of cellular subpopulations or models,” said Tim Jackson, PhD, Senior Image individual cells.” Processing Engineer, Sartorius BioAnalytics Product Development. “Researchers now have an unprecedented, high-quality label-free segmentation resource and starting point for training neural networks. Microbioz India, September 2021 Edition | 22

Featured Article The high-throughput Incucyte® system was Due to the nature of neural network-based essential for building the image dataset as it allowed algorithms being orders of magnitude more capture of a very high volume of high-quality complex than traditional image analysis, this data images. Use of a high-throughput label-free culture set will allow for more robust segmentation of system eliminated the risk of biological artifacts, various cell morphologies, and ultimately minimize while leads to increased confidence in the output of user-introduced biases.” algorithms based on the dataset. Prior to launch of the LIVECell dataset, the largest Source: Sartorius Newsroom dataset of label-free images available to researchers For further information on the LIVECell data consisted of 4,600 images derived from 26,000 set, please visit: https://sartorius- cells. research.github.io/LIVECell/ For more information on the Incucyte® Sartorius collaborated with the German Research system, please visit: www.sartorius.com/incucyte Center for Artificial Intelligence (DFKI) to demonstrate the utility of this dataset and plans to continue work with the Center to further advancements in deep learning for the life sciences community. Images of the eight different cell lines (human breast cancer (3), human glioblastoma, human hepatocyte carcinoma, human neuroblastoma, human ovarian cancer, mouse microglia) were captured every four hours, over the course of three to five days using an Incucyte® Live-Cell Analysis system. Microbioz India, September 2021 Edition | 23

Featured Article Detection of Mastitis Pathogens in Raw Milk of Symptomatic Cows Mycoplasma and especially Mycoplasma bovis as well Staphylococcus aureus and Streptococcus This application note describes a method for the agalastiae are harmful pathogens for cows detection of different pathogens with the Mastit 4C and calves, which can cause Kit by DNA Diagnostic on the qTOWER³ and inflammation and the Mastitis disease. Mastitis leads proves its specificity. to a reduction in the quantity of milk, which, in addition to animal health and welfare, is also an Materials and Methods important economic factor for farmers. Inveterinary The Mastit 4C Kit from DNA Diagnostic wasused to diagnostics it is important to isolate the harmful isolate bacterial DNA from milk. The number of bacteria from the milk and to detect them precisely. samples of the protocol was adapted. All steps were Since performed in a 1.5 ml reaction tube. After the transfer of the Pre-lysis buffer from the 96 Mycoplasma bovis has no cell wall and is therefore Deep Well Plate to the 1.5 ml reaction tube, 500 µl of naturally resistant to antibiotic therapy, therapy is raw milk were added. One sample was contaminated particularly difficult. Only isolation of the with different pathogens to test the performance of symptomatic cows can prevent it fromspreading. the extraction and detection. The extraction of bacterial DNA from milk is a challenge due to the diversity of ingredients, such as fat, sugarand proteins. That’swhyaspecialextraction kit or procedure is recommended, for instance the Mastit 4C Kit by DNA Diagnostic which is perfectly optimized for the extraction of bacterial DNA from milk. The extracted nucleic acid can then be detected qTOWER³ Series using the qTOWER³ for quantitative real-time PCR and the Mastit 4C Kit from DNA Diagnostic. To detect Mycoplasma, Staphylococcus and The detection kit Mastit 4C Kit detects up to four Streptococcus in the samples, the detection part of targets in parallel like Staphylococcus aureus, Streptococcus the Mastit 4C kit of DNA Diagnostic in combination agalastiae and Mycoplasma bovis and Mycoplasma species and with the qTOWER³ by Analytik Jena were used. The provides an amplification control in addition. qPCR protocol was adapted to the qTOWER³. Microbioz India, September 2021 Edition | 24

Featured Article Samples and Reagents Sample 5 shows Ct-values for HEX (Staphylococcus  Mastit 4C Kit (DNA Diagnostic) aureus), ROX (Streptococcus agalactiae) as well for Cy5  Raw Milk (Dairy farm) (Mycoplasma species). Instrumentation Conclusion  qTOWER³ (Analytik Jena) The results of the IAC show, that the amplification of  qTOWER³ Software: qPCR soft3.4 the Mastit 4C Kit in Analytik Jena’s qTOWER³ is working perfectly together. First of all, we resume, Results and Discussion that the whole experiment is valid due tothe results of Before the values were checked for the different the NTC (No template control) and Positive control. pathogens, the internal amplification control was Furthermore, the Internal Amplification Control is applied to verify the results. The criteria for valid present in all samples so that results of the different results is an Internal Amplification Control (IAC) raw milk samples from cows with symptoms of lower thana Ct-value of 32. Mastitis are positive or negative for the several All samples show Ct-values between 23 and 31 so pathogens. To summarize this experiment the the result of the other targets are acceptable. combination of qTOWER³ (Analytik Jena) and Mastit 4C kit (DNA Diagnostic) could be a solution Samples number 2 and 3 show positive IAC, but are for customers, which want to detect pathogens in negative for all the other channels, resulting in a milk. negative diagnosis. Samples number 1 and 4 show a For more information: positive signal for the ROX channel, resulting in [email protected] contamination with Streptococcus agalactiae, whereas www.analytik-jena.com the signal for sample number 4 is very low. Microbioz India, September 2021 Edition | 25

Featured Article Environment Research & Heavy Metals: A Toxic Relationship What are heavy metals? This potential exposure can then be put into a Heavy metals are naturally occurring elements & toxicological perspective by comparison with the are a group of metals and metalloids that have so-called provisional tolerable weekly intake relatively high density and are toxic even at ppb (PTWI) values for toxic metals, which have been levels. Examples include Pb, As, Hg, Cd, Zn, Ag, established by the Food and Agriculture Cu, Fe, Cr, Ni, Pd, and Pt. Organization of the World Health Organization (FAO-WHO) (De Smet, 1999; WHO, 1981, 1979). Multiple industrial, domestic, agricultural, medical Many pharmacopoeias contain heavy metal and technological applications have led to their detection methods that are usually based on color wide distribution in the environment; raising reactions with special reagents such as concerns over their potential effects on human thioacetamide or diethyldithiocarbamate, and the health and the environment. amount present is estimated by comparison with a Their toxicity depends on several factors including standard (WHO, 1988a). the dose, route of exposure, and chemical species, Why carry out heavy metals testing and as well as the age, gender, genetics, and nutritional analysis? status of exposed individuals. Interestingly, small background levels of these These metallic elements are considered systemic elements are common in the environment but large toxicants that are known to induce multiple organ amounts of any of them may cause acute or chronic damage, even at lower levels of exposure. They are toxicity (poisoning). This is why regular heavy also classified as human carcinogens (known or metals testing and analysis especially within probable) according to the U.S. Environmental industry is so important. Protection Agency, and the International Agency for Research on Cancer. This review provides an Heavy metal toxicity can result in damaged or analysis of their environmental occurrence, reduced mental and central nervous function, lower production and use, potential for human exposure, energy levels, and damage to blood composition, and molecular mechanisms of toxicity, lungs, kidneys, liver, and other vital organs. genotoxicity, and carcinogenicity. The potential intake of the toxic metal can be estimated on the basis of the level of its presence in the product and the recommended or estimated dosage of the product. Microbioz India, September 2021 Edition | 26

Featured Article Hence Abdos has come up with the idea of Maxi Long-term exposure may result in slowly RCF Trace metal-free centrifuge tubes progressing physical, muscular, and that are free of 18 different Trace metals. neurological degenerative processes that mimic Alzheimer’s disease, Parkinson’s  Abdos maxiRCFTM Trace Metal disease, muscular dystrophy, and multiple Free centrifuge tubes are sclerosis. Allergies are not uncommon and repeated long-term contact with manufactured using high purity some metals or their compounds may virgin USP Class VI Medical grade even cause cancer. polypropylene, conforming to US Environmentally Relevant Most FDA 21 CFR, free from natural Hazardous HMs and Metalloids rubber & heavy metal. Heavy metals are among the most  Abdos maxiRCFTM Trace Metal investigated environmental pollutants. Almost any heavy metal and metalloid Free Centrifuge Tubes withstand may be potentially toxic to biota 20,000xg RCF. depending upon the dose and duration of exposure. Many elements are classified  Pure, Additive Free, Natural into the category of heavy metals, but Color Cap for easy labelling and some are relevant in the environmental context. Trace Metal free.  Abdos maxiRCFTM Trace Metal List of the environmentally relevant most toxic Free Centrifuge Tubes work heavy metals and metalloids contains Cr, Ni, Cu, Zn, Cd, Pb, Hg, and As. excellently in high density gradient Heavy metal pollutants most common in the separation of samples environment are Cr, Mn, Ni, Cu, Zn, Cd, and Pb. In 2009, Many reports has suggested four metals,  Abdos MaxiRCFTM Trace Metal i.e., Cr, Cd, Pb, Hg, and the metalloid As, as the Free centrifuge tubes are highest priority pollutants for control. Some other heavy metals are also hazardous to living organisms specifically manufactured to depending upon dose and duration of exposure. ensure that common trace metals For example, Mansouri et al. have found Ag as more toxic than Hg to a freshwater fish. and elements that can How Important Is Trace Metals Analysis in interfere with experiments are Your Specimen Samples? below an ICP-MS detection level We at Abdos realized the fact that collection and of one part per billion (< 1ppb). transport of your samples have to be organized in Metals included in testing are: Trace metal-free collection lab ware. And if a specimen is received in a non-metal-free container, Zinc, Iron, Copper, Manganese, abnormal results or false positives will be reported. Aluminum, Silicon, Nickel, Vanadium, Phosphorous, Cobalt, Chromium, Lithium, Selenium, Cadmium, Mercury, Arsenic and Lead. Scan for more details Author Name: Vinod Joshi Product Manager - International Div., Abdos Labtech Pvt ltd Microbioz India, September 2021 Edition | 27

Business News IBM Research and Arctoris Thomas A. Fleming, Arctoris Co-Founder & accelerate closed loop COO, explained “The future of drug discovery is drug discovery with AI and computational, with AI and robotics paving the Cloud way for better treatments to reach patients sooner. We are excited about partnering with IBM Source: IBM Research Research on a world-first closed loop drug discovery project bringing together two leaders in I BM Research and Arctoris announced they the field of AI and robotics-powered drug are investigating the application of AI and discovery. This collaboration will showcase how automation to accelerate closed loop the combination of our unique technology molecule discovery. IBM Research has platforms will lead to accelerated research based on better data enabling better decisions.” developed RXN for Chemistry, an online platform “This collaboration is a great example of the leveraging state-of-the-art Natural Language enablement that AI, Cloud and Automation can have in the space of material design. The Processing (NLP) architectures to automate integration between the two complementary technologies reveals how it is more and more synthetic chemistry. Representing chemical important in R&D to turn great research into great reactions via SMILES (Simplified Molecular Input viable products.” said Dr Teodoro Laino, Distinguished Scientist at IBM Research Europe – Line Entry System), the system is able Zurich. to perform highly accurate reaction Dr Matteo Manica, Research Scientist at IBM predictions using its powerful AI. Research Europe – Zurich, coordinating the Optimised synthetic routes are then project, stated that: “This is a unique opportunity to quantify the impact of AI and automation used as input for RoboRXN, an technologies in accelerating scientific discovery. In automated platform for molecule our collaboration, we demonstrate a pipeline to perform iterative design cycles where generative synthesis. models suggest candidates that are synthesized with RoboRXN and screened with Ulysses. The data Arctoris has developed Ulysses, an produced by Ulysses will then be used to establish end-to-end automated platform for a feedback loop to retrain the generative AI and improve the proposed leads in a completely data- drug discovery research. The platform driven fashion.” ensures accuracy, precision, and The collaboration is currently on-going; financial terms were not disclosed. reproducibility by leveraging robotic experiment execution and digital data capture technologies across cell and molecular biology and biochemistry/biophysics. Experiments conducted with Ulysses generate more than 100 times more datapoints per assay compared to industry standard, leading to deeper insights and accelerated progress compared to manual methods. The two platforms are now being combined for the first time in a research collaboration that will see new small molecule inhibitors for undisclosed targets being designed, made, tested, and analysed (DMTA) in an autonomous, closed loop approach. Concretely, IBM Research will design and synthesize novel chemical matter (Design, Make), to be profiled and evaluated by Arctoris (Test, Analyze), with the resulting data informing the subsequent iteration of the DMTA cycle. Microbioz India, September 2021 Edition | 28

Microbioz India, September 2021 Edition | 29

Business News Existing approaches to predict developability and identify sequence liabilities that are based on ENPICOM introduces a simple sequence analysis scale well in terms of powerful liability prediction the required computational resources, but do not solution to de-risk antibody development provide sufficient accuracy, as they fail to consider the 3D structure of the antibody. Source: ENPICOM Structural modeling, on the other hand, provides detailed insight into the surface exposure of E NPICOM, an innovative unwanted motifs, leading to highly reliable bioinformatics software company, liability predictions. Performing these announced today at the Biologics UK predictions in a high-throughput manner is conference new IGX Platform capabilities to accurately annotate exposed liabilities, perform incredibly challenging and time-consuming, as it structural modeling of antibodies at scale, and requires significant computational resources and determine developability profiles using specialized software infrastructure. ENPICOM customizable penalties. This major development solves these challenges by integrating SAbPred allows researchers to improve their candidate directly into the IGX Platform. SAbPred is selection by making accurate developability developed by the Oxford Protein Informatics predictions for thousands of sequences at the Group and is a validated, peer-reviewed, and same time, in a secure, intuitive environment. globally recognized toolbox for accurate and “The spectacular progress in the field of antibody efficient structural analysis of antibodies. discovery calls for more powerful tools to “The three main challenges in the analysis of effectively leverage all generated data. liabilities we aim to address are accuracy, ENPICOM has tackled the challenge of throughput, and ease of use.” commented Dr candidate pool expansion with the launch of the Nicola Bonzanni, co-founder and Chief Product Antibody Discovery Module earlier this year. The new liability analysis capabilities we are adding Officer at ENPICOM. “The IGX Platform today will enable scientists to easily select better enables researchers to fully utilize the added antibodies with even greater confidence”, said value of NGS data. You can accurately predict Jos Lunenberg, co-founder and Chief Executive exposed liabilities for thousands of sequences at Officer at ENPICOM. once, easily assess the developability of From thousands of sequences to that needle-in- candidates, and proceed with your selection with a-haystack antibody. confidence.” Feature highlights The new IGX-Annotate App enables scientists to accurately predict exposed liabilities, configure custom liability penalties, and integrate developability information throughout their entire workflow. In addition, researchers can now better gauge antibody developability by comparing characteristics of newly discovered antibody candidates to those already brought to the market. The new Select page provides a flexible environment to compare and rank the most promising antibody candidates based on user- defined characteristics. It brings antibody sequences, experimental metadata, and in silico predictions together into one comprehensive overview, making it easy to select the best candidates to express in the lab or move further down the development pipeline. Microbioz India, September 2021 Edition | 30

Business News Scientists using the IGX Platform can now: • Use a validated tool to independently perform structural modeling analysis and accurately annotate liabilities. • Increase candidate success by flagging exposed liabilities early in the discovery phase and identify antibodies with the best developability properties. • Annotate structural liabilities for thousands of sequences and integrate the developability information throughout their entire workflow. • Configure their own liability penalties through The first issue of PRONEWS focuses on the purpose and passion that drives the team at an intuitive UI to compute scores that align with APCC. The first story in this issue of their de-risking strategy. PRONEWS is about personalized medicine alias • Benchmark candidate developability profiles precision medicine – which is a treatment of against a database of clinically validated cancer at its molecular profile. Precision antibodies. Oncology is all set to revolutionize the way • Overlay developability characteristics on Cancer is diagnosed, monitored and treated. information-rich visualizations like phylogenetic The other stories describe the medical efficiencies of the medical physics team and the trees to prioritize and select the best candidates. pediatric cancer combined management team. The IGX Platform and the Antibody Discovery The issue also has three patient case studies Module provide an end-to-end cloud solution addressing various types of cancers such as that enables researchers to easily perform Cytoreductive Surgery and HIPEC gynecologic complicated tasks such as clustering, phylogeny, cancer, proton therapy for breast cancer and the and prediction of exposed liabilities in a secure evolving landscape of systemic management of lung cancer. and scalable environment. With the addition of Commenting on the occasion of the launch, Dr these new tools, researchers can now expand Prathap C Reddy, Chairman of Apollo Hospitals, their candidate pool and improve their antibody said, “Technology has percolated every industry, selection. and one must keep up with the trends to offer To learn more about performing accurate liability best patient care. From the latest technologies predictions and de-risking antibody development and cancer treatments, we’re now revolutionizing with the IGX Platform, join the free webinar on the way we update our patients, clinicians, partners and followers about new initiatives and September 28, 2021. the latest developments in cancer care and at APCC, on a regular basis. It gives me great Apollo Proton Cancer pleasure in launching the India’s first video Centre Launches India’s newsletter- PRONEWS by Apollo Proton First Video Newsletter Cancer Centre.” Source: AP Sharing her thoughts, Dr Sapna Nangia, Editor in Chief of PRONEWS Apollo Proton Cancer Cancer Centre (APCC), the first Proton Centre, said, “At Apollo Proton Cancer Centre, Therapy Centre in South Asia & the we tread new ground as the first center in this Middle East, launched India’s first video part of the world to offer proton therapy. newsletter – PRONEWS. Aimed at reaching readers with the latest in Cancer care and treatment, PRONEWS will be a quarterly compilation that will educate, empower and elevate the knowledge of its readers. Microbioz India, September 2021 Edition | 31

Business News It’s such an honor to contribute to various life science fields from cardiovascular research to We continue our leadership with site specialized robotic surgery and immunology and even groups working with personalized medicine. We also offer organoids.” – said Kjell Steps, Account Manager comprehensive cancer care comprising at CytoSMART Technologies cancer surgery, chemotherapy and image guided intensity modulated radiotherapy, The CytoSMART Research Grant in a nutshell: administered with compassion and 1.The applicant can apply for a Lux2, Lux3 FL, diligence. The new format of our or Omni live-cell imaging microscope. newsletter, then, naturally, reflects this 2.The applicant will be provided with the pioneering spirit. It is my pleasure, as the editor-in-chief, to be present at the necessary training needed for operating the launch of the first video PRONEWS, a device. capsule depicting our passion for and 3.The digital microscope can be used for 4-18 expertise in cancer management.” months. 4.The applicant must be currently registered at an After a successful winter educational institution or employee of a research initiative, CytoSMART group or research institution. Research Grant project resumes again 5.The applicant must perform research that will highly benefit from using real-time imaging Source: CytoSMART Technologies devices. A minimum qualification of an undergraduate degree. CytoSMART Technologies today opened To learn more about the CytoSMART Research the application of CytoSMART Research Grant and application process, visit the official Grant for 2021 Autumn. The CytoSMART Research Grant is an initiative to CytoSMART website. help motivated students and researchers improve their project’s quality. The grant offers one of the About CytoSMART CytoSMART live-cell imaging devices that can be CytoSMART Technologies is a specialist in the used for research purposes for a specified period. development and manufacturing of smart The CytoSMART automated incubator-friendly microscope systems for life science laboratories. microscopes are suitable for monitoring cell The company was founded in 2012 by a team of cultures in any research topic that could benefit biologists and engineers who were convinced from live-cell imaging. The application period starts on September 6 and will last until October that a new generation of miniaturized 15, 2021. microscopes, powered by artificial intelligence “We started this initiative at the beginning of for image analysis, would allow biologists to 2021, and we never thought it could be as make discoveries more efficiently and at scale. In successful as I turned out to be. The number of 2018 CytoSMART was selected by Microsoft for applicants and the innovativeness of their its prestigious Scale Up program. CytoSMART’s research proposals were outstanding. Also, the fact that live-cell imaging devices can play such a valuable role in these projects, is an indescribable reward for all the efforts we have put into these systems. microscopy solutions are used in research laboratories worldwide. Microbioz India, September 2021 Edition | 32

Business News Viedoc Technologies achieves certification to ISO/IEC 27001:2013 Source: Viedoc Technologies He continues, the ISO certification will benefit our clients by speeding up the auditing process and Viedoc Technologies was recently assuring them of the highest standards in information presented with its ISO/IEC 27001:2013 security, critical in clinical studies. certificate affirming that the company’s Requiring accreditation of vendors is an increasing operation globally, adheres to the trend today. And Viedoc is one of only a few in the internationally recognized standard for industry to fully meet this requirement. The ISO/IEC information security management. The certification certification assures new clients of the highest quality ensures clients worldwide that Viedoc has and level of data security when using Viedoc. implemented processes and procedures for This certificatin is a huge competitive advantage and implementing, maintaining, and continually takes us closer to our ambition of being one of the improving its information security management leading eClinical Suite players in the market, adds system. Mats. We have always been committed to following a high- About Videoc quality and consistent security management system to guarantee all data we process is well protected. But to Viedoc Technologies is an ISO 27001 certified developer of ensure we are doing our best and make it easier for modern cloud-based eClinical systems, founded in Uppsala, our clients to easily verify this, we set out to achieve Sweden. Since its inception in 2003, Viedoc is challenging the the ISO 27001 certification, says Jens Pettersson, standard for conducting clinical trials by creating user-friendly, Chief Information Security Officer. scalable software that complies with the demands of every phase The certification process has enabled us to adopt and of a clinical study – making what matters more accessible. Our implement the applicable requirements for an commitment to working for a healthier world by accelerating all information security management system (ISMS) at aspects of a clinical trial brings therapies to the market faster. As a Viedoc. It includes people, processes, and IT systems result, Viedoc is trusted by 9 of the top 10 pharmaceutical by applying a risk management process, adds Jens. companies, as well as leading research institutions and businesses Obtaining this certification is a recognition and within Biotech, CRO, Medical Device and Consumer health, all promise to our clients that our processes are world- over the world. Visit www.viedoc.com to learn how your Clinical class, says Mats Klaar, CEO. Study can benefit from Viedoc’s eClinical Suite. About the certificate ISO/IEC 27001 is an international standard on information security management. It is known worldwide for providing requirements for an information security management system (ISMS) Microbioz India, September 2021 Edition | 33

Business News IB1001 has a very compelling safety profile, easy oral administration [sachet mixed with water], affirming its very IntraBio Reports favourable risk/benefit profile as a treatment for this Statistically Significant and devastating disease.” Clinically Meaningful Professor Antony Galione, FRS, FMedSci, Improvements in the Use Statutory Professor of Pharmacology, University of of IB1001 for Treatment of Oxford commented, “GM2 Gangliosidosis (Tay- GM2 Gangliosidosis (Tay- Sachs and Sandhoff disease) is a devastating disease Sachs and Sandhoff that has never had any available treatment. We are disease) very excited that IB1001 is the first drug that is effective for this disorder and will improve the lives Source: IntraBio of so many patients and their families. Given what is known about IB1001’s mechanism, and its IntraBio Inc today announced positive data multiple successful clinical trials, we will continue from its multinational clinical trial of IB1001 to investigate this drug for other rare genetic (N-acetyl-L-leucine) for the treatment of neurological diseases and for more common GM2 Gangliosidosis (Tay-Sachs and Sandhoff neurodegenerative diseases prevalent in society disease). with large unmet medical needs.” IB1001 demonstrated a statistically significant and In a joint statement, Rick Karl, President of the clinically meaningful improvement in symptoms, Cure Tay-Sachs Foundation and Dan Lewi, Chief functioning, and quality of life in both the primary and secondary endpoints for pediatric and adult Executive Officer of the Cure Action for Tay-Sachs patients with GM2 Gangliosidosis. Foundation, commented: “This treatment is a The trial met its primary endpoint, the Clinical Impression of Change in Severity (CI-CS), which major breakthrough for the GM2 Gangliosidosis was assessed by blinded, centralized raters community that includes Tay-Sachs and Sandhoff. (professors of neurology with expertise in It is the first drug to offer hope to the patients and movement and neurological disorders). The trial also met secondary endpoints including families affected by these devastating diseases. the Scale for the Assessment and They are progressive, life-threatening conditions Rating of Ataxia (SARA), the with no approved medicinal treatments. There is Modified Disability Rating Scale (mDRS), the Investigators’, an urgent need for this effective treatment to be Caregivers’, and Patients’ approved and made available for patients in our Clinical Global Impression of Change (CGI-C) assessment. community before the window of therapeutic IB1001 was observed to be safe opportunity is lost.” and well-tolerated, with no drug- related serious adverse events. “The results of this study are hugely important for the GM2 community,” said Dr. Susanne Schneider, Principal Investigator and Professor of Neurology from Ludwig Maximilian University of Munich. “IB1001 is the first drug to demonstrate a statistically significant and a clinically meaningful effect for the treatment of GM2 Gangliosidosis. Microbioz India, September 2021 Edition | 34

Business News The positive results of this IB1001-202 study are reinforced by the efficacy and safety profile of IB1001 already demonstrated in IntraBio’s successful IB1001- 201 study for Niemann-Pick disease Type C (NPC). As is the case in the IB1001-202 clinical trial for GM2 Gangliosidosis, IB1001-201 was the first clinical trial to demonstrate statistical significance and a clinically meaningful effect in patients with NPC. These results provide further momentum for the broad clinical development program the employees of Sakura. They are the key to the planned for IB1001 which will address high unmet strong brand Sakura Finetek Europe is today. I will medical needs for treatment of both rare and focus on extending our solid culture of innovation, teamwork, and customer centricity to be able to common neurological disorders. empower pathology professionals, now and in the Mateo Iglesias Vice future.’ Chris Koeman, General Manager at Sakura President Marketing & Finetek Europe: ‘We look forward working together Sales Sakura Finetek with Mateo. We believe he has the experience and drive to Europe enable the ambitious plans Sakura has. His progressive and human approach is what makes him the ideal candidate to SSource: Sakura Finetek help us bring Sakura to the next level.’ akura Finetek Europe appointed Mr. Mateo More about Sakura Finetek Europe Iglesias as Vice President of Marketing & Sales EMEAI. Mateo starts September 1st At Sakura Finetek Europe we advance cancer and will be responsible for all marketing and sales diagnostics by providing integrated solutions for activities for Sakura’s fast-growing markets. anatomic pathology and patients through best-in- Mateo has over 20 years of experience with class innovations, quality and customer care. This (bio)medical companies. For the last fifteen years, is how Sakura became the original manufacturer of he worked for Agilent Technologies, as Associate Tissue-Tek®, Cyto-Tek®, Tissue-Tek VIP® and Vice President EMEAI. His proven leadership Tissue-Tek Xpress®, today’s industry standards. capabilities and marketing & sales experience will be a massive benefit in this new role. With headquarters based in Alphen aan den Rijn Focus on people and culture (NL) and seven regional locations in Europe we The new Vice President is honoured and thrilled of empower pathology professionals in the whole his appointment and has a clear vision: ‘I believe in EMEAI region. For more information visit www.sakura.eu, follow us on LinkedIn, or contact us via [email protected]. Microbioz India, September 2021 Edition | 35

Product Showcase Silicone tubing especially designed for dairy industry! Milk contributes more than 60 per cent Our Platinum Cured transparent silicone Tubing of the total livestock production and known for its ultra-smooth bore to prevent any accounts for nearly 67.69 per cent of particle entrapment, reduces the chances of the value of the output of the livestock sector’s butterfat occurrence and it can also help to contribution to the national GDP. To alleviate eliminate the probability of Microbial growth the labour involved in milking, much of the within the milk transfer line and ensures smooth milking process has been automated but still they transfer of milk. face many challenges during milk processing in dairy Industry which leads to loss of money, loss Features and Benefits of time, loss of quality and much more. 1. Superior Flexibility, Kink Resistance First and foremost, kink resistance is very properties. important - In Dairy Industry kink resistance 2. Clear Transparency allows easy visual properties plays a vital role as kinking can happen Inspection of Milk during process. when a cow stucks in tubing or it moved around 3. Smooth bore to ensure less during Installation which can cause leaking and contamination and resists butterfat build-up. loss of milk - A huge messss!!! 4. Water repellent properties allow easy cleaning and complete soakaway. Secondly, Tubing must be weather resistance so 5. Excellent Weather resistance properties. that it doesn’t become stiff especially during 6. Non-toxic, non-haemolytic. winter season as stiffness can lead to delay in 7. Free of plasticizers like production since it takes much time and becomes phthalate/Bisphenol-A very difficult during installation. 8. Higher Shelf life Also Tubing should be flexible but still retains its 9. Free of any heavy metals and hazardous elasticity of feeding retention and not become substances. too easily stretched out. Additionally, tubing 10. Sterilisable by using different techniques must have tear resistance ability as it can be like autoclaving, Ethylene oxide, Gamma torned on / off feedings, dropped on floor or a radiation. cow steps on it that’s why tubing with tear 11. Dimension range 1mm ID – 3000mm resistance ability very crucial to avoid constant ID dimensionally accurate & excellent repair. workmanship and finish It also must have resistance to milk fat 12. Customized Colour, dimension and absorption which can attract dirt and negatively hardness available on demand. infect hygiene as milk fat build-up softens the 13. Good flex fatigue resistance enhances tubing and reduces its strength or even cause Milking peristaltic pump life & Milking discoloration. Dosing Pump Life. After examining all this challenges and to overcome it, we at Ami Polymer Pvt Ltd come- Major Applications up with one stop solution and here we are Introducing Platinum Cured Silicone Tubing 1. For Raw milk transfer. which ensures milk processing or transfer of milk 2. For Bulk milk transfer. must be safe, hygienic and high performance. 3. For Rotary Milking Parlour line. Platinum cured silicone Tubing is designed for 4. For Milking Dosing Pumps. the application in Automatic milking system / 5. For Milking Peristaltic Pumps. rotary milking parlour/ robotic milking / Peristaltic Pumps. Microbioz India, September 2021 Edition | 36

Product Showcase Physical Properties. Authored by: Simran Singh Compliances Designation – Sr. Executive International Sales & Marketing Manufactured and packaged in clean room of class About Amipolymer Pvt Ltd: 10000 facility audited by TUV Nord. FDA 21 CFR 177. 2600 AMI POLYMER PVT. LTD., Established in 1998 at Silvassa in USP CLASS VI Dadra and Nagar Haveli, is leading Exporter of silicone tubes, TSE/BSE FREE silicone gaskets in India. AMI POLYMER PVT. LTD. is one of Trade PHATHALATE FREE India's verified and trusted sellers of listed products. With their ROHS COMPLIANCE extensive experience of supplying and trading Platinum Cured DMF #26201 Silicone Bi Color Tube/ Color Coded, AMI POLYMER PVT. LTD. has made a reputed name for themselves in the market with high \"The Company believes in “Be in business with quality Platinum Cured Silicone Bi Color Tube/ Color Coded, ethics” and assure Quick Development, Silicone Corks & Stopper etc. Focusing on a customer centric Fastest Delivery, Ultimate Quality and also very approach, AMI POLYMER PVT. LTD. has a Pan-India presence Competitive Rates\" and caters to a huge consumer base throughout the country. Buy silicone tubes in bulk from AMI POLYMER PVT. LTD. at Trade Visit us at www.amipolymer.com Take a sneak-peak look at our India quality-assured products. advanced facility AV https://www.youtube.com/watch?v=V51mA1nOYQM. Feel the best ever purchasing experience with Ami Polymer Pvt Ltd- GET An OFFER NOW! Email: [email protected] / [email protected] Microbioz India, September 2021 Edition | 37

Product Showcase All You Need to Know About the Zebrafish Model in Biomedical and Applied Research Zebrafish (Danio rerio) are increasingly used Predation pressure led to the development to address key questions raised in basic of shoaling behaviour, believed to reduce and applied research including, but not stress and aggression among fish held in limited to, biomedicine, toxicology, small groups. Of course, while conditions environmental science, biotechnology and in the wild can vary considerably, attempts aquaculture.1 Zebrafish have proven well-suited have been made to provide stable to large-scale genetic screening, whether in husbandry conditions in laboratory search of models for human disease or for settings. It is important to emphasise that when control mechanisms related to natural or pathological differentiation of cells and tissues, implementing lessons from fish in their natural studies which are more difficult to carry out in habitats, some caution is warranted: laboratory mammalian model organisms because of their strains have been maintained in research facility long in-utero gestation periods. aquaria for at least 25 years (c. 100 generations). A long list of established zebrafish models aids in understanding the mechanisms behind diseases, During this time, diverse adaptations and and novel toxicology models have been domestication steps have arisen, for example developed for endocrine disruptor studies. Many concerning mechanisms of sex determination. In clinically relevant biochemical pathways are addition, a vast quantity of internet-based conserved between fish and human, but cannot resources is available to aid both beginners and be reliably investigated in vitro, in unicellular more experienced researchers using the zebrafish organisms, or in invertebrates. model (zfin.org). Worldwide, more than 1000 laboratories use zebrafish as a research model (zfin.org). In Europe, the tightly interconnected zebrafish scientific community consists of more than 350 laboratories (eufishbiomed.eu) that would benefit from guidelines for standardised husbandry conditions at the European/international level. The guidelines provided here are meant to constitute a framework for European zebrafish research groups, and a basis and reference point for future welfare relevant studies. The biology of zebrafish Zebrafish are native to South Asia where they are found in streams, ponds and other slow-flowing water bodies including rice fields. The range of environmental conditions under which zebrafish live includes water bodies with temperatures ranging from below 10℃ up to 40℃, clear or turbid water, pH from 6 to just below 10, conductivities from 10 to 271 µS/cm, depth from surface level down to 60 cm, a wide range of bottom substrates (primarily silt), presence or absence of vegetation and elevations ranging from sea level to over 1500 m altitude (Figure 1). Microbioz India, September 2021 Edition | 38

Product Showcase Figure 1. Temperatures and pH levels measured Two recent international surveys on zebrafish at 35 natural zebrafish habitats at altitudes husbandry practices from over 100 facilities in between 14 m and 1576 m above sea level (blue five continents showed that parameters like dots). Ranges recommended for zebrafish physical housing, nutrition, pathogen status, housing systems (pH 6.5–8 and 24–29℃; green water conditions and holding densities can be area) and values commonly referred to in highly variable. literature being optimal for reproduction (pH In addition, international zebrafish resource 7.4–7.5 and 28℃; red circle) are indicated. centres and several single facilities have recently published husbandry procedures that lay the The Directive 2010/63/EU groundwork for educated discussions for a The 3Rs (Refinement, Reduction and gradual standardisation of zebrafish protocols. Replacement) are an intrinsic part of the EU Although less common in earlier literature, there Directive 2010/63 on the protection of animals is now an increasing trend towards including used for scientific purposes and apply to all non- detailed information about the husbandry human vertebrate animals, including parameters used in studies in the Materials and independently feeding larval forms (Article 1). Methods section of publications. With the notion However, fish are not elaborated on and specifics that parameters like feeding, temperature, dark– for zebrafish are not provided at all. This light period etc. may affect results from motivates the work on common European experimental research, it is recommended to guidelines for ensuring sound husbandry and always include a detailed description of the high standards of welfare for zebrafish. zebrafish husbandry parameters used. Important A key issue within the Directive is the time point factors for dissemination of experiences and when fish larvae can be regarded as skills are national/regional/international training independently feeding and free-living, thus initiatives with theoretical and hands-on courses. falling under the scope of the legislation. The The major application of Zebrafish in development of poikilothermic animals is Biomedical Research involved in: temperature dependent, and for zebrafish this Recommendations for housing and critical time point is 120 hours post fertilisation husbandry (hpf) at 28.5℃. The guidelines provided here address and suggest International exchange of experience and recommendations for: transportation and best practices reception, safe introduction into facilities, During the last few years, dedicated husbandry housing systems and environmental conditions sessions have become mandatory at international (temperature, dark–light cycle, water quality, zebrafish conferences.6 stocking density, environmental enrichment, feeding), breeding, identification and nomenclature as well as sanitisation of equipment. View larger version Transportation and reception Exchange of fish between laboratories usually poses three main challenges: organising safe shipment of fish, ensuring compliance to fish welfare as well as to national and international legislations, and finally avoiding spreading pathogens between laboratories. For genetically modified zebrafish lines, regulations related to a transgenic or genetically modified organisms (GMO) biosafety and restrictions must also be fulfilled. If available, documentation of health monitoring and relevant husbandry parameters used for the originator population should precede the shipment. Microbioz India, September 2021 Edition | 39

Product Showcase Stocking density Embryos hatch around 60 hpf and settle over the Quarantine for biosecure introduction of bottom of the Petri dish/tank until roughly 5 new strains/fish days post fertilisation (dpf) when the swim After safe reception of embryos or fish, the next bladder has developed sufficiently to allow challenge is to avoid the spread of potential swimming. Body mass, rather than numbers of pathogens into the main section of a facility. animals, sets the limits for fish/l density recommendations. Thus, the density of juveniles While egg surface disinfection (bleaching) can be higher than the corresponding figures strongly improves biosecurity, it does not suffice given for adults. to safeguard against dissemination of intracellular Environmental enrichment pathogens. It is strongly recommended to keep In general, observing normal behaviour patterns and breed newly imported animals under and the absence of any signs of illness or stress quarantine containment to limit the risk of among the fish suggests that well-being is pathogen dissemination into the main facility. maintained. Two commonly used laboratory Water and housing systems measures for animal welfare are reproductive Decades of experience have proven that success and plasma (or water) cortisol levels. maintaining zebrafish is a relatively Changes in the D–L cycle, water conditions (e.g. straightforward task. In most cases, commercially rise in ammonia levels) or presence of parasites available tank systems are used, which come fully (e.g. nematodes) are associated with reduced egg integrated with filter systems, germicidal production. irradiation (UVC) and light and temperature Feeding control units. It is generally accepted that a combination of live Temperature feeds and processed dry feeds improves growth, Fish are poikilothermic and the zebrafish is a generation time and reproductive performance, tropical species (although many natural habitats all positive indications of well-being. Dry feed are temperate, Figure 1). The commonly used diets are generally assumed to be nutritionally references to developmental stages relate to hpf complete, whereas live feed and the associated determined by the speed of development at fish prey-capture behaviour have an enrichment 28.5℃. Temperature influences water chemistry effect. Formulated diets simplify feed delivery, and animal physiology. storage and preparation. Dark–light cycle Breeding, identification, nomenclature In nature, light conditions vary with seasons and Among the attributes that make the zebrafish a weather. Modern laboratory facilities present the great model species is its high fecundity. A female fish with a static dark–light (D–L) cycle can spawn hundreds of eggs per single mating. It (commonly 10 hours dark, 14 hours light, is however recommended to set up fish for optionally with gradual decrease and increase in spawning with a recovery period of at least one light intensity mimicking dusk and sunrise). week to allow for sufficient regeneration and Water quality maturation of new ova. Depending on the local water supply used for Sanitisation of equipment and hygiene recirculating water systems, some laboratories A clean environment is essential for maintaining can use tap water without major amendments. a high standard of animal health and welfare. Chlorine still needs to be removed, as levels safe for humans (c. 0.1 mg/l) are toxic for fish. For more information: Constant stirring and aerating of the water for 24 Girish Nair hours prior to use is sufficient to remove Founder, Director chlorine, whereas water sanitised with Biolinx India Pvt Ltd chloramine (which is less volatile) needs to be [email protected] filtered through active charcoal. In most facilities, the water chemistry needs to be adjusted before use. Microbioz India, September 2021 Edition | 40

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Product Showcase High-Throughput Mannose Glycans Screening of Crude and Purified mAb and Non-mAb Protein Sample – Sartorius Octet® GlyM Kit You can Download application note on Mannose Glycans Content Screening for Human-mAb Samples Using the Octet® GlyM Kit. G lycosylation, an important post- Reliable data at this early stage supports shorter translational modification, is a key lead times and prevent later failures. Assay critical quality attribute (CQA) that Workflow the GlyM Kit utilizes mannose specific lectin which is immobilized onto the biosensor influences product safety and efficacy and should surface (Figure 2). This surface binds to mannose structures in the sample. be monitored when developing new biological The secondary amplification steps increase the drugs. Among various glycan structures, mannose binding signal from protein of interest (POI) and negates the minimal signal from host cell proteins is especially important as it affects a protein’s (HCPs). This increases assay sensitivity, facilitating the analysis of crude cell culture samples without plasma the need for purification or enzymatic digestion. This essentially eliminates the need for any sample clearance and preparation beyond a dilution step, simplifying the mannose screening workflow for users. reduces its in Additionally, the GlyM Kit reduces time to results and increases sample analysis throughput. As a vivo half-life, result, 96 clones can be screened in under 1 hour on the Octet® RH96 system. negatively impacting drug efficacy. While process conditions, media and feed formulations are critical factors in the optimization of protein mannosylation, optimal cell line selection can be a limiting step; early screening for mannose content can facilitate the selection of clones that produce proteins with favorable glycan profiles during cell line development (CLD). The Octet® GlyM Kit enables high-throughput mannose content screening in crude cell culture samples during early clone selection stages in the CLD process. When combined with titer analysis, it expedites the selection of the top performing cell lines producing proteins with the most favorable glycan (mannose) profile (Figure 1). Only clones which are the most suitable will move downstream for further optimization. Microbioz India, September 2021 Edition | 42

Product Showcase The GlyM Kit comes with the biosensors and The Octet® GlyM Kit is designed to provide cell reagents required to screen mannose content in up line development and upstream bioprocessing to 96 samples and is compatible with Octet® groups with a ready to use kit for the rapid relative systems with at least 4 channels. Relative Mannose mannose content screening of both purified and Content Screening the GlyM Kit enables the cell harvest antibody samples. Unlike traditional screening and ranking of crude samples based on techniques, the Octet® platform can also monitor the relative mannose content in the POI. The kit other functional critical quality attributes (CQAs). provides ranking results for samples with mannose content that differs by 4% or more (Figure 3, see The ability to view and choose from desired titer samples with 3% vs. 8% and 10% vs. 14% Mannose and mannose levels at the same time provides more content). Users can assign all screened clone in-depth knowledge that facilitates more informed samples into groups depending on the mannose decisions in the early selection of optimal clones content (e.g., high, medium or low) and focus during cell line development.In this application further development effort on the desired group of note, a novel approach for relative mannose clones only. Data Analysis Octet® Analysis Studio content screening and ranking of human software combines titer data with mannose content monoclonal antibody (mAb) samples is showcased; data for more in-depth analysis that facilitates using a well characterized mAb from the National selection of the best clones. Institute of Standards and Technology (NIST) as well as samples developed at Novartis. Source: Octet RED384 System (sartorius.com) Mannose Glycans Content Screening for Human- mAb Samples Using the Octet® GlyM Kit Microbioz India, September 2021 Edition | 43

Pharma News Dr Reddy’s gets DCGI gesture to lead stage 3 trial of Sputnik Light in India Dr Reddy’s Laboratories was granted approval by the drugs regulator to conduct Phase III trials of single- dose COVID-19 vaccination Sputnik Light. According to Hindustan Times, the subject expert committee (SEC), of the Central Drugs Standard Control Organisation CDSO (CDSO), had recommended to the Drugs Controller General of India(DCGI) that approval be granted to conduct Bridging Studies in India in order to support Sputnik Light. RDIF has partnered up with Dr. Reddy’s Labs in order to distribute the first Trials in the United States at that time were 250,000,000 doses of the Russian vaccination paused on reports of rare blood clots. and conduct the necessary trials in India for regulatory approval. The drugmaker’s withdrawal comes as India In July, the Subject Expert Committee of tackles legal challenges with manufacturers over CDSCO had denied authorisation for emergency indemnity issues, with its junior health minister use to Sputnik-Light. This ruling ruled out the saying last week that a team had been formed to engage with vaccine makers. necessity of conducting the phase III Russian “This team is in continuous dialogue with Pfizer, vaccine trial in the country. Moderna and Johnson & Johnson to discuss and address India says Johnson and various issues including the issue of indemnity,” Bharati Johnson pulls out proposal Pravin Pawar had said. Johnson & Johnson and the Central Drugs for quick COVID-19 vaccine Standard Control Organisation (CDSCO) did not immediately respond to Reuters’ requests for approval comments on why the company withdrew its application. Source: Reuters As of July 31, Johnson & Johnson is yet to request a full approval for its shot with the U.S. India’s drug regulator said on Monday that FDA, while Pfizer Inc , BioNTech SE, and Johnson & Johnson withdrew its proposal Moderna Inc have already sought full approval of seeking accelerated approval of its COVID- their vaccines with the FDA. 19 vaccine in the country, without giving In India, the drug regulator had given emergency additional details. use authorisation to Moderna’s vaccine in June. The U.S.-based company had said in April it was seeking an approval to conduct a bridging clinical study of its Janssen COVID-19 vaccine candidate in India. Microbioz India, September 2021 Edition | 44

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Pharma News Aleor Dermaceuticals gets USFDA gesture for generic antibacterial gel Source: PTI Granules India and Jubilant Cadista Recall Drug firm Alembic Products in US Market Pharmaceuticals on Monday said its joint venture Aleor Source: PTI Dermaceuticals has received approval from the Granules India and Jubilant Cadista, two generic US health regulator for generic Metronidazole drug manufacturers, are recalling several goods in gel, used in the treatment of inflammatory lesions the United States, the world’s largest market for of rosacea. medicines, for various reasons. According to the Aleor Dermaceuticals Ltd has received final latest Enforcement Report issued by the US approval from the US Food & Drug Food and Drug Administration (USFDA), Administration (USFDA) for its abbreviated new Granules India’s US-based unit is recalling over drug application (ANDA) for Metronidazole gel 1.14 crore Naproxen Sodium tablets, a non- USP, 1 per cent, Alembic Pharma said in a steroidal anti-inflammatory drug used to treat regulatory filing. pain, menstrual cramps, and inflammatory The product is a generic version of Galderma diseases such as rheumatoid arthritis, gout and Laboratories LP”s Metrogel gel 1 per cent, it fever. added.According to IQVIA, Metronidazole Gel, USP 1 per cent, has an estimated market size of USD 29 million for twelve months ending June 2021, Alembic Pharma said. The product is indicated for the topical treatment of inflammatory lesions of rosacea. Aleor had previously received tentative approval for this ANDA, it added. The company has a cumulative total of 149 ANDA approvals (132 final approvals and 17 tentative approvals) from USFDA, Alembic Pharma said. Shares of Alembic Pharmaceuticals were trading at Rs 783 per scrip on BSE, up 1.10 per cent from its previous close. PTI AKT BAL Microbioz India, September 2021 Edition | 46

Pharma News According to USFDA, the company’s US-based Aurobindo Pharma gets USFDA gesture for unit is recalling the affected lot due to ‘Current malignancy treatment Good Manufacturing Practice (CGMP) injection deviations’. Source: PTI The affected lot has been manufactured at Drug firm Aurobindo Pharma on Friday Granules India’s Telangana-based plant. The lot said its arm Eugia Pharma Specialities has been distributed in the US by New Jersey- has received approval from the US based Granules USA, Inc, the report stated. health regulator for Cyclophosphamide injection, used in the treatment of several types of cancer. Granules initiated the Class II voluntary recall on The company”s subsidiary “Eugia Pharma 11th August this year. Specialities Ltd has received a 505(b)(2) NDA approval from the US Food & Drug As per USFDA, a Class II recall is initiated in a Administration (USFDA) for its Cyclophosphamide Injection 500 mg/2.5 mL situation in which use of, or exposure to, a and 1 g/5 mL vial,” Aurobindo Pharma said in a violative product may cause temporary or regulatory filing. The product will be available in ready-to-use medically reversible adverse health consequences injection preparation, it added. “The product shall be launched in the US market or where the probability of serious adverse health in the near term and will be manufactured in consequences is remote. Eugia”s manufacturing facility in India,” USFDA further said US-based Jubilant Cadista Aurobindo Pharma said. Pharmaceuticals Inc is recalling 14,544 bottles of Donepezil HCL Tablets, which is used to treat mild to moderate dementia in Alzheimer’s disease. The company is recalling the affected lot due to it being ‘subpotent,’ it added. Jubilant Cadista Pharmaceuticals is a fully-owned subsidiary of Cadista Holdings Inc, which is a part of the Noida-based Jubilant Life Sciences Company. The affected lot has been produced by Jubilant Generics in India and marketed in the US by Salisbury-based Jubilant Cadista Pharmaceuticals, Inc. Jubilant commenced the Class III recall in the US on 20th August 2021. As per USFDA, a Class III recall is initiated in a ‘situation in which use of, or exposure to, a violative product is not likely to cause adverse health consequences.’ According to industry estimates, the US generic drug market was estimated to be around USD 115.2 billion in 2019. It is the largest market for pharmaceutical products. Microbioz India, September 2021 Edition | 47

Pharma News According to IQVIA, the approved product has an addressable market size of USD 170 million for the twelve months ending June 2021, it added. Cyclophosphamide Injection is indicated for malignant lymphomas, multiple myeloma, leukemia, mycosis fungoides, neuroblastoma, adenocarcinoma of the ovary, retinoblastoma and carcinoma of the breast, the filing said. This is the 19th product to be approved (including 3 tentative approvals) for Eugia. Aurobindo now has a total For this purpose, a joint venture company shall of 482 abbreviated new drug application be incorporated with an aim to enter the (ANDA) approvals (453 final approvals and 29 respiratory biosimilars space. tentative approvals) from the USFDA, it added. Shares of Aurobindo Pharma were trading at Rs Under the terms of the agreement, the joint 709 per scrip on BSE, up 1.56 per cent from its venture will leverage Cipla and Kemwell’s previous close. PTI AKT BAL complementary strengths for end-to-end product BAL development, clinical development, regulatory filings, manufacturing and commercialisation of Cipla and Kemwell biopharmceutical products. Cipla’s respiratory Biopharma announce prowess combined with Kemwell’s expertise in execution of a joint venture biologics will accelerate bringing these essential agreement for developing, products to market. Umang Vohra, Managing Director & Global manufacturing and Chief Executive Officer of Cipla, said: “This commercialising development reinforces our strength as leaders in biosimilars respiratory therapy and is in line with our aspiration of global lung leadership. Guided by our purpose of ‘Caring Source: Cipla for life’ our partnership with Kemwell will help serve unmet needs of patients across the globe.” Cipla Limited (BSE: 500087 | NSE: Anurag Bagaria, Chairman and Chief Executive CIPLA EQ, hereafter referred to as Officer of Kemwell, said: “We are very excited “Cipla”), a leading global pharmaceutical about combining the strengths of Cipla’s company and Kemwell Biopharma Private expertise in development and commercialisation Limited, a leading biopharmaceutical Contract of respiratory products and Kemwell’s biologics Development and Manufacturing Organisation capabilities. Through this JV, we aim to make (CDMO), announced execution of a joint cost-effective biotherapeutics available to more venture agreement to develop, manufacture and patients globally.” commercialise biosimilars for global markets. Microbioz India, September 2021 Edition | 48

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