GENE TALK

[00:00:48]Good evening and sir.

[00:00:50]>> Good evening.

[00:00:51]>> Good evening sir.

[00:01:04]Sir, >> good evening. Good evening.

[00:01:07]Good evening, sir.

[00:01:10]>> Not sir. I'm a student.

[00:01:12]Don't say sir.

[00:01:15]I'm I'm learning so much from you. So I'm a student.

[00:01:20]>> Okay. Okay. Okay.

[00:01:23]>> Very old students.

[00:01:51]Omar Raj Kamar Raj Kumar doesn't attend your talks sir that I'm very angry with him he's busy with his own work sir >> become nervous when he is thereh no sir Good evening Dr. Roshan.

[00:02:19]>> Uh hello ma'am. I also try and share my s >> if you want you can try.

[00:02:26]>> Uh good evening sir. Good evening sir.

[00:02:29]Good evening sir.

[00:02:33]>> Good evening. showing host uh has disabled the participant screen sharing.

[00:02:38]I think I'll also have to be a co-host.

[00:02:41]>> Oh, okay. Okay.

[00:02:43]>> Good evening. Madam, good evening.

[00:02:45]>> Good evening. Good evening to Wanda sir, good evening.

[00:03:18]>> Good evening.

[00:03:25]I think it was.

[00:03:45]>> Good evening. Nice to see you. Jenist.

[00:03:50]>> Good evening, Shankar G. Sir namaste sir.

[00:03:55]>> No sir. No sir. Student sir don't make that mistake again.

[00:04:00]>> No sir. You are you are my teacher. You are teacher of uma and you are teacher of all teachers.

[00:04:06]>> No no no no. That may be a past story. I am now presently a student.

[00:04:13]>> So so now your two students are our close geneticist. Um as well as praa both are same match.

[00:04:25]Good evening Shanda sir.

[00:04:27]>> Good evening. Good evening.

[00:04:30]Good evening.

[00:04:33]>> Good evening from and yourself.

[00:04:38]>> Good evening.

[00:04:42]>> Good evening. Good evening. Good evening Shang.

[00:04:46]Good evening sir. Good evening.

[00:04:49]>> Good evening. Good evening Gopy. We can start.

[00:04:55]>> Uh we can start.

[00:04:57]>> Yeah.

[00:05:00]>> Start start. We'll start.

[00:05:05]>> Thank you. So um good evening to one and all and it's my pleasure and privilege to um welcome you all to this uh genetic webinar and this time we have um two eminent speakers two different topics and uh this is our third talk on the series gene talks the first talk is on inborn of metabolism all of you know it's a uh individually rare but collectively common disorder and to talk on this we have a very eminent speaker Umam Mahishi madam she is the professor in HOD uh SRMC I mean department of neonatlogy SRMC Chennai and the second talk is on genetic testing based on cytogenetics and molecular uh cytogenics by Dr. Roshan he is the chief consultant and the HOD of Kim's hospital strvand and uh um we have these talks are one is it's entirely different talks entirely different topics but um I think they are connected and coming to my um uh duty today I would like to welcome uh the president Kerala Dr. Nandmar Sar for his um unwavering support to all the academic programs by the genetics chapter.

[00:06:33]>> Thank you. Thank you.

[00:06:34]>> And also welcome to I I would also like to welcome Dr. Gobi Mo secretary AP Kerala and Dr. Binotan treasurer all the officials of AAP Kerala and also Dr. Jos is the uh vice president's house central and all the seniors like um um Parvati Madame Shanger sir um go sorry um Mahinda sir and um lot of seniors are there push sir everybody is there so welcome to this webinar and I also like to welcome the speakers today Dr. Madam and Dr. Roshan to this gathering and I'd also like to welcome all the delegates all the students and all the uh members of AAP to this gathering.

[00:07:38]So I'm sure that this um um today's academic feast will contribute to the practicing pediatricians as well as uh the students of um genetics as well as postgraduate students and uh and I would call upon Nandumar Sar for his presidential address.

[00:08:03]Thank you.

[00:08:04]>> Yeah. Good evening respected Dr. Girish person of the genetic chapter secretary Dr. Vinita and Dr. Jo south vice president CIP Dr. Gopi Dr. Bin Kerala and today's speakers Dr. Mohammed Shari and Dr. Joshin Daniel and most respected senior teachers Parodi Madam Puramasar Sinasanar Dr. Shanka Moendas and others teachers on this platform my dear friends and my dear students. So at the let me at the outset let me genetic chapter for regularly conducting very useful for practicing piations as well as our students topics which is genetics which we are rarely used to expose all this and very difficult to understand while reading but someone teaches us definitely a good thing for as a practice immigration to learn how to proceed and how to suspect and how to uh evaluate a case of and when to rougher and all these cases. As you all know, genetic has moved from being a niche specialty to become an integral part of everyday pediatric practice. With advances in genetic genomic medicine, many conditions that once remained un undiagnosed can now diagnose early, enabling precise diagnosis, targeted treatment and informed family counseling and in some cases even prevention. From newborn screening to development disorder to metabolic diseases, rare syndromes and precision therapeutics, genetics is transforming the way we care for children. So to discuss two important topics about immigration and Daniel and I wish all the very best for this program happy learning thank you all over to Dr. Girish.

[00:10:01]So I will introduce uh um madam and then we'll straight away go to the um talk.

[00:10:11]Okay.

[00:10:17]So introducing Dr. Omar Mahir. Madam actually she doesn't need any introduction. Uh she's a well-known speaker and she's a professor in NHOD uh department of neonatlogy. SRMC uh Chennai and she has lot of awards and publications in to her credit including uh more than 50 papers in index journals and uh she has authored many chapters in books and peer-reviewed national and international journals and she is the research ambassador of SRNC and she is currently the executive member of AP Neoch and Sonel AP coordinate for the perinatlogy module also she's a trainer NRP national level trainer. So over to you madam.

[00:11:09]I will and unshare it. Okay. Okay. Thank you.

[00:11:21]>> Good evening all. Thank you for the kind introduction and thank you for the opportunity given to me to talk on this uh gene talk. Uh like let me see how I could uh you know do my job. So my topic is on approach to IM. Um so this is uh the spectrum of metabolic disorders which we have seen in the rare disease clinic in a 2-year period. Um so what the spectrum of disorder tells us is whatever we used to read in Nelson book um like asm we are seeing in the clinic that is because of increased improved availability diagnostic availability you can see the tag which comes as like incidence is rare but though the incidences are rare individually they are collectively common that is why we could see the spectrum and also we need to remember that uh this phase of uh we are like more consanguinity exist in India and also some endogamus population is there uh so that is why IM comes across as a major uh you know chunk of disease and also I would put uh like amongst genetic disorders which cannot be treated many of these IM are treatable so these are our metabolic stars I request to mute someone has these are our metabolic stats which we see in the clinic. So my objective I have broken my approach into two different objectives. First one is to when to suspect IM and how to evaluate and manage. So when to suspect IM as I said it might be rare but collectively common. So uh people might think that IM should be considered only when there is a positive family history. It is not. So first it is majority autotosomal resis where the parents are going to be carrier and they are going to be normal.

[00:13:28]U you have to suspect whenever there is a consuity even if they are non-anszy married it can be an endogamous population or married within uh you know that particular area. So we do get without any positive family history. So this one has to remember that is the importance of putting a three generation pedigree whenever we are evaluating a child with suspected IM. So IM mimic sepsis is very much true. IM mimics sepsis but consider IM workup uh when you think that there is no risk factor for sepsis and the baby is presenting poorly with the features of shock and metabolic acidosis. Um and sometimes IM can also coexist with sepsis like um E.coli with galactosemia, Canada sepsis with multiple caroxil deficiency.

[00:14:22]Remember to uh you know difficult to remember each presentation. I feel so uh you know the complex metabolic pathways might be difficult to remember but what we need to use is algorithmic approach rather than um you know kind of knowing each and every pathway and also we have to uh you know from a broad screening test we can go to high highly specified test in subsequent slides I will show you how to do that. Um some might think that you know only if you are treatable why should we evaluate etc. There are two important aspect to it. One is mainly are treatable. It can be a very simple treatment like giving a biotin for biotinidase deficiency. Also for the parents who thought that the child antiatal period period was absolutely fine. They got discharged home and why is the baby again getting admitted in NICU or even in PICU in later stages. We have to give an answer to them. There can be unexplained something unexplained developmental delay etc. which can present. So for formal genetic counseling also we need to evaluate.

[00:15:36]Some might think even we used to think once upon a time that TMS is diagnostic.

[00:15:42]Um so it is even if the baby has a normal newborn screening if the baby present symptomatic or they have had a TMS test you have to you know considering re-evaluation and the am testing does not stop with TMS alone.

[00:15:59]So when we come to disorders as such it can be protein disorders which includes amino acidopathies organic acidia and uriocycle disorder. These presence as intoxication that means a metabolites gets accumulated as it gets accumulated the presentation or the severity worsens. Carbs and lipids are mostly energy produces. So they present with energy failure or energy production problem. So it can be gluconioenesis or glycogen storage disorder. Third disorders includes fatty acid oxidation defect or cholesterol synthesis defect. So clinical presentation if we have to divide very simply into group one, group two and group three disorder. Group one disorder is intoxication. They present with poor feeding lethargic and distress or seizures after a initial symptom free period. their antiatal scans are absolutely fine and the baby presents later in life with these kind of features. The classical example would include nalapurine disease or um metal melanic academia, isovaleric academia or propionic academia or disorders of ura.

[00:17:08]Group two they present because energy failure. So in in case of prolonged fasting or infection or stress can make the child have presentation. It can be acute. It can present in newborn itself or it can be chronic. Whichever uh organs involves more energy, require more energy in terms of heart, liver, muscle, brain. So they the they are the one which present with energy failure.

[00:17:35]Group three is a broader perspective. It includes both storage disorders as well as vitamin response disorders. It could include peroxyal disorders with Zelve as the prototype or it could include lysosomal storage disorders and vitamin responsive disorders. So it is partly treatable only group one and group two disorder where fatty acid oxidation defect can be treatable. Group three disorder by and large storage disorder.

[00:18:05]Some have enzyme displacement but the group three also includes vitamin responsive disorder which is easily treatable.

[00:18:14]So clinical presentation wise they can present with enkopathy seial hypotenia or liver dysfunction, liver failure, choleisteris, cardiac failure or arithmia. Other presentations include the hypoglycemia um like antinatal health syndrome or sometimes catact or dysmorphisms they can present with presentation like per oral ulceration and alopeia and sepsis like picture. So these are like categories as as such as I said group one is intoxication group two is energy failure group three is storage and vitamin responsive disorders. So presentation of minion can be remembered very well like acute enkapilopathy, bacterial sepsis like picture, C for cardiomyopathy, D for dysmorphism, death unexplained or dermatological, enlarged liver, family history, jaundice, high drops or impractable hiccups.

[00:19:11]So objective two, how do we evaluate and manage? So for that like you know sub we know that substrate is getting converted into a product. If that is absent so you have this um the substrate which gets accumulated or the substrate gets accumulated in u into it kind of you know travels one more pathway and this metabolite can also be toxic. So toxic accumulation is one thing and or there can be deficiency in the product which results in energy failure. So by and large your diagnostic evaluation would include either evaluating these substate which is getting increased or this metabolite which is getting increased or the product which is low. So this enzyme or co-actor is we know that the gene is responsible for coding the protein. So it can be evaluated in molecular also with the availability of molecular testing.

[00:20:10]Sometimes there could be a reverse phenotype which is happening. They would have got evaluated and the gene diagnosis would have come, genetic diagnosis would have come first and then we go back to the biochemical testing.

[00:20:21]Though the norm is do the biochemical evaluate and then go for genetic evaluation. So it can be summarized as GA which stands for glucose, AG, lactate, ammonia and ketones. These are the metabolites which can be done very easily in many of the labs plus specialized test which includes TMS, urine GCMS in terms of detecting urine organic acid, HLC for plasma amino acids and enzyme like biodenidase deficiency or g enzyme etc. So it can be very well remembered by gall. Most of my residents when they have a suspected IM they report saying that GA is normal or GA is showing glucose is low and ketones low etc. So it's a very uh simple pneumonic which anyone can remember and evaluate and of course we could do a molecular analysis after as a chair too. So case scenarios are always interesting. Let us go through some of the case scenarios to understand the approach better. So this is a term baby presented on day 17 with sh cry dais with the berubin of 25 enkopathy. The baby also has vomiting and poor weight gain. Even though the working diagnosis is acute beline enkopathy we would have seen any number of neonates with 25 berubini they do not present on day 17 and they do not behave the way this baby behaved. And also there was deterioration in the sensorium and needing ventilation. There is no blood group incompatibility and the M MR showed basil ganglia hyperintensity.

[00:22:03]So somehow the team suspected IM this is the MRA which showed the basil ganglia hyperintensity.

[00:22:12]So when we did the gallop we found that there was acidosis and there was hyperammonia with the ammonia of 289 with the ketones being positive. So definitely the baby's pouring of ketones and ammonia is high and also there is acidosis. So what do we do? We take the algorithm and see where it fits into. So this baby has neonated hyper ammonia and the symptoms happen after 24 hours and there is acidosis. So that points towards organic academia. So investigation showed one is high. One is also a very simple investigation to say whether we are kind of dealing with organic esthemia's ideology for hyperammonia or ura cycle disorder as ideology for hyperammonia because one will be low in ura cycle disorders because ura is not produced. This might be this might also give us an clue towards diagnosis of hyperammonia.

[00:23:13]TMS showed uh you know the elevation of certain metabolites sgest of either a methyl melaninic or propionic academia.

[00:23:22]Plasma amino acid showed a elevated glyin and urine GCMS showed an elevated hydroxy propionic academia. So don't get perplexed with this slide. It just simply says that you will have a the the investigations they would have given you the report. So all the report it's it comes to the diagnosis of propionic academia. So this is a quick card for the postgraduate students over here with the gak approach how do we come to the diagnosis. So when there is an acidosis with hyperammonia and ketones positive it suggest organic acidia. When there is a very high ammonia with respiratory alkyossis, it suggest ura cycle disorder. When there is a hypoctootic hypoglycemia, it suggest fatty acid oxidation defect.

[00:24:13]So the other ideology could be due to hyperinsulinism. But fatty acid oxidation disorder generally do not need that much high compared to insulin hyperinsulinism.

[00:24:25]So other could be increased lactic per se which can uh you know present in fibroid dehydrogenase or pyroate caroxilase deficiency. Other the thing can be gala normal which with the glucose ABG lactate ammonia normal with ketones alone elevated suggestive of maple syrup urine disease. So in this baby as we know that there was acidosis with hyperammia with ketones it suggest organic esthemia. So the molecular also confirmed the diagnosis of propionic academia though and had multiple crisis managed with special formula and breast milk and this is the baby one year. The treatment modality is very simple. So this is uh this is one of the slides which I like the most. So you can decrease the substrate availability or you can provide some diffusion metabolites or you can try to increase the enzyme activity or you increase the disposal of these toxic metabolites. If we follow these four simple principles of management you can apply this in any and you can get the treatment done. So ma if you have apply the management principles in treating propionic hmia what do we do? We keep NPO initially followed by some breast milk as to reduce the substrate availability. So also subsequently we have to give low protein diet in terms of vegetarian diet. Also metronidazol can be given for 7 days in a month to reduce the chain fatty acids which is again a source of propionic acid. So decreasing substrate availability. Um so then increasing toxic disposal in a acute state can be hemmoiialysis or hemopiltration. Ideally if you do not have provision you can do a peron diialysis you can do a carnitine supplementation to increase the toxic disposal. So increasing enzyme activity in topionic academia is by biotin supplementation. What is the pearl you learn? You get an ammonia level when there is an something unexplained there but helps in differentiating and also follow the management principles. Moving on to the case to firstborn of third degree consignas couple discharged on day three presented on day nine with irritability abnormal movements. Um so there was lip smacking hyperonia very poor sensorium but a was normal. So we did a gak and also tabc temperature airway breathing and circulation was maintained. Of course sexis can be a great mimicker. So antibiotics was also started. The main principles would be to promote anabolism and to prevent catabolism and remove toxics.

[00:27:16]So what what was done was done that gala is normal. Ketones alone was positive.

[00:27:23]So the baseline investigation showed ketone. Now we know that we can you know go to a diagnosis of MSUD as such. Um but how do we confirm? We have to send it TMS and urine GCMS. But as we know not all the labs have facility to do we might have to send it outside. What is the quicker way to diagnose it? We can do a MRA with MRS. MRS would show a peak in branch chain amino acid level and MRA would show a double swan appearance. You can see that this is one span and this is another swan. So the changes in the midbrain cerebral pedle and the dentritates of the cerebellum leads onto this double swan appearance which could be considered typical of MSUD. So what do we do? Take this uh management principle and apply. We did nail per nail protein for 24 to 48 hours followed by quantified amount of breast milk along with special formula. Diamond supplementation can be done and diialysis or hemodialysis or peronial diialysis can be done. So high energy is very important. Lipic can be started after reasonable exclusion of fatty acid oxication. If there is hypoglycemia let it be. Do not use your G. Instead start on insulin whose other ICU management is also as important as supportive care is also important as specific management. So nail perural how long this is some of the thing which we face as you know clinician many times I have seen that they never get restarted on breast milk or they kept milk peroral for a very long period which itself becomes catabolic because the muscle mass gets you know lied and the protein gets the baby becomes catabolic and the protein or the ammonia levels get increased because of the catabolism itself. So what is what we should do? Protein should not be restricted for more than 24 to 48 hours. Even when the baby is on diialysis, we can start some trophic feeds. How we start in newborn period for pre-term etc. And also the paragamantra is if that is working use the gut. So you do not want a trophy of the gut etc. So give some quantified amount of breast milk. How we manage in our unit is we procure this specific formula called as a AMD1 formula and keep it in pharmacy. So um what happens this particular formula it does not contain any protein. It has high kind of carbohydrate and fat and high energy. So it can be used for pretty much for any of the protein disorders. It can be used for MSUD. It can be used for propionic academia. It can be used for methyl melanin. So once we start on this because we cannot procure all the formulas and keep in the pharmacy. So on discharge it can be replaced with MSUD specific formula which is available from Restine which is Indian based company.

[00:30:37]So the misconception which is here is breastfeeding is contraindicated. Many of many of the pediatricians might think uh like this but this milk contains you know it has very good natural protein source and all amino acid levels are lower and it should be started as early as possible after the crisis. So if you compare here the human milk with the cow's milk all the amino acid levels are lower in human milk. So it can be a very safe alternative for natural proteins.

[00:31:09]So here the loin isolucin came as th00and and we managed same very similar also we did the car supplementation. Um, so this is a baby getting Contrast difference between the two baby they are one and the same. So this is in the ICU where the baby is arching and having a fistonous portion. This is a baby in the clinic. Same baby who has come out of crisis. Um so we we should always think optimistic on treatment of metabolic. We should not consider okay metabolic we cannot treat.

[00:32:05]So, IM and diet will be like a big combo. So, you have to kind of manage both together and there are ways to do it. Um like you know breast milk along with some formula. So what we do is quantified amount of breast milk along with special formula. There are other food also for older children. So this is a follow-up chart which is done. So here you can see from 2,00 it has come down to less than 500. Ideal level is to bring it less than 200 even though less than 400 of loin is um acceptable. So apart from that is there any other methodology to treat? Can you increase the enzyme activity by any other method?

[00:32:50]Yes, we do have modality for liver transplantation. So this particular baby was sponsored for liver transplantation as well. For large molecule disease you give enzyme from outside. These are small molecule disease. So you transplant the liver. In the transplanted liver this enzyme activity will be present. So that will serve the purpose. For some of them it is curative. For some of them it can kind of resolve the crisis. It will keep them under control etc. For organic academia as such it is a partial cure. For us for ura cycle disorder, MSUD etc. You can consider liver transplant as a full cure. So we have to optimize nutrition and assist the neurology. And four important question which which we have to ask before liver transplant is is IM potentially curable? uh is the severity of the IM in terms of recurrent metabolic crisis or poor quality of life warrants liver transplant because liver transplant is not an easy option. So you have you have to also ensure that whether all other systems are preserved especially the neurology and also do the parents give consent after counseling and also explaining the risk versus benefit ratio. So what is the p you learn from this? Not to keep nail perural for longer than 24 to 48 hours.

[00:34:13]What is the importance of breast milk and also monitor amino acid level and dietary management. So moving on to case three. This is a firstborn of non-conus couple presented uh you know referred as protein metabolic disorder. Gallup showed hypoglycemia, hyperamonymia and ketone strays. So admission weight that baby had severe failure to thrive. So um because they thought that it is a protein disorder they have stopped breast milk and they have started on this particular AMD1 diet. Whenever they reintroduced breast milk the ammonia elevated so they stopped again they put on a AMD1.

[00:34:55]So when clinically examined we found absent red reflex capital but sensorium was normal stool color was normal urine was not showing any type of staining. So we know that clinical examination is very important. So the total galactose was normal because the baby was on nil you know the AMD1 does not have any lactose. The diet enzyme was very low suggestive of galactosemia.

[00:35:22]to how to explain the normal galactose because the baby's already on galactose free diet. Though the the referred physician did not know that they are indirectly putting on lactosefree diet because they started on this AMD1 diet because it contains one thing only malttodextine automatically you know the baby was on nil lactose.

[00:35:48]So in view of clinical suspicion we started on galactose preform formula we did the followup counseling also especially like you know regarding the endogenous production and also premature ovarian failure in a girl baby. So this is a molecular testing which showed the calin positive. So what is the pearl you've learned? Clinical examination is very important. Cat look for red reflex.

[00:36:13]Time and again we say that hy examination you should never forget to have the carrier of a red reflex in all hyperonymia could be due to liver dysfunction. This baby had failure to fly because of UTI and also on not the correct appropriate formula. So moving on to group two disorders. So this is a baby who had transient hypoglyasmia during neonatal period but she was not evaluated a third degree consignment couple presented later in life around two two and a half year old with seizures and kapopathy he had hypoglycemia and continued to have seizures treated developed like deep coma with tonic posturing and no evidence of menitis or toxic exposure.

[00:37:00]So the treating physician in fact gave a call saying that there is tonic forine.

[00:37:05]Should we withdraw car but when we evaluated g it showed a low glucose with ammonia mildly elevated 127. So 100 we can take it as normal ketones with a hyper you know lipmic serum. So what we followed? We followed the BMDG guidelines and the TMS was suggestive of one of the fatty acid oxidation defect.

[00:37:30]So what is the treatment modality for this particular fatty acid oxidation defect as we restricted protein for protein disorders? We have to restrict protein or modify the protein in lipid sorry we have to restrict the lipid or modify the lipid in lipiders and also we have to provide adequate calories. So diialysis can be done in the initial stages glucose corn starch or special formula can be given to provide efficient metabolites. We can give four co four factors like visa fibbrate which is available as an adult formulation. We can use alternate substrate like triptinino but it is very costly. So we have to use carnitine here with very cautiously we have to use not in all longchain fatty acids or carnitine cannot be used for all the metabolic disorders as such. So even the like treatment recommendation in this particular workshop they kind of they had the consensus um to abandon in particular intravenous carnitine during metabolic derangement.

[00:38:39]So how long they can fast neonate can fast up to 3 hours maximum they can fast up to 4 hours. So this is a followup of the same child who is on restricted fat educated about sick day regime and she's on regular followup like including echo heart. So she is the girl who is now having like you know um entering into adolescence.

[00:39:04]So emergency management we have to remember that should be started at the first sign of illness because hypoglycemia is a late event. Uh so management wise avoid fasting avoid fasting and avoid fasting. So what we have to take home here is carnitine you you need to use very judiciously.

[00:39:27]Hypoglycemia is a late event in fatty acid oxidation defect. Whenever there is a symptomatology like vomiting etc itself the baby should be admitted and put the dextrose fluid and increase the dextrose concentration because you do not want them to go on to hypoglycemia do not wait for hypoglycemia to treat the group three disorders um which are vitamin which would include vitamin responsive disorder also among group two disorders these are treatable.

[00:39:57]We had this particular baby who got referred from another tertiary care hospital on day eight to us was P2 L2 non-consuous marriage term baby normal transition discharge on day three. So baby had seizures from day five and we got the baby on day eight of life g is normal till this point for all the case scenarios gak is helping us. There are certain neurotoxicity syndrome which we call it as the neurotoxicity type two disorders where glet can be normal. So what happened is like the baby was hemodynamically stable. There was no seesures but you know the baby's tone was reduced. On day two of admission, baby developed multiple episodes of seizures and on probing the history um the mother clearly told that she had increased fetal movement because it was gravid or two she could say that she perceived increased fetal movement. So we started on mazolum infusion followed by pyodoxin well controlled after the next day itself. So the MRI showed T2 hyper inensity, MRS showed lactate PE and PET showed burst suppression pattern.

[00:41:13]We went ahead with the investigation.

[00:41:16]These are specialized investigations.

[00:41:18]Till now we have been learning about TMS and urine GCMS. There are certain special investigations which called this called as pipolic acid and AACA that was elevated in this particular baby suggestive of pyoxin dependency issues.

[00:41:33]Of course we did the molecular which showed problem in the antiquity gene.

[00:41:38]So this is the serial pics of the same baby who presented with refractory seizures and had birth suppression pattern with white matter changes. A child is developing normally. Um so developmentally normal uh even they come for followup regularly even though they have moved to approach. So here what is the availability I would like to highlight that you know IV preparation of pyrooxin is not available as standalone but it is available as neurocine gold and the pyrooxil phosphate which is an active form is available as tetra plus.

[00:42:18]So what is the pearl you learn from here? History taking skill is important.

[00:42:22]Give a trial of pyroin refractory seizures. So again as a neonitologist I would like to uh you know put forward to all of you here that newborn screening is lifesaving. It is a preventive measure. So here is a clinical presentation of biotin deficiency with alopecia and skin rash. Here is a baby who got diagnosed by new screening itself in our hospital. So he's doing perfectly normal. So newborn screening helps. There are variable resource various resources available. One is British inherited metabolic disease group which is called as BMDG. So there are emergency guidelines available. So we do give print out of these emergency guidelines when they get admitted. So there is another tool in the same BMG called as temple tool which is tool enabling metabolic parents learning. It is a very simple PPT format which a parent can learn even a beginner can learn using this tool.

[00:43:22]So there are various resource material available. Um so this particular book sod is a beautiful book comprising of all the inborn errors of metabolism in a very descriptive manner. I'm I'm sure all of you know Dr. Anel Jalan who also has authored this inborn error of metabolism in critically ill newborn um and I am also humbled to author this genetic and inherited metabolic disorder in children. So this is again one more e resources available very metabolic and this is also a beautiful e version. So moving on to echo message anything unexplained please think of like our first scenario we thought burbin and kflopathy then turned out to be propionic acidmia um so there could be the protein metabolic disorder they thought hypermonia protein metabolic disorder but turned out to be galactomia so we have to you know be vigilant thorough and systematic history taking skills is important and also a thorough head to foot examination is also important. You can start evaluating the GA followed by TMS and urine GCMS.

[00:44:40]Occasionally the GA is normal. You might have to resume to specialized testing need to follow flow chart and we also learned about simple principles of management in terms of decreasing substate etc. treatment is often dictated by experience very I want to insist to this forum that breastfeeding is not contraindicated in all do not stop breastfeeding completely with the exception of galactosemia or infantal care all others we have to quantify the amount of breast milk and give along with specialized formula in fact ura disorders and organic asmia especially the distal ura disorder they uh you know tolerate breast milk very well. Team effort and holistic approach is a key.

[00:45:32]Thank you.

[00:45:41]Uh thank you. Thank you madam for the excellent talk on uh approach to inoness of metabolism. you made this uh approach very simple and lucid and highlighting the importance of the subtle clues in the history and physical examination. So we'll take up the uh questions at the end of the second talk. We'll move on to the um second talk.

[00:46:16]Okay. So, Binita can you share the slide of u >> seen >> that slide visible? Yeah.

[00:46:52]>> Okay. Ah yeah. Okay. It's visible. So uh the next talk is on um cytogenetics and molecular cytogenetics and we have Dr. Roshan Daniel and he's the head of department of medical genetics Kim's health hospitals trwandram Kerala and he has uh qualifications European board certification and PGD precision oncology and has publications over um more than 15 articles in index journals and his interest is in pediatric rare diseases and adult rare diseases and um um over to you Dr. Russian.

[00:47:38]>> Thank you, sir. I'll share my screen.

[00:47:42]>> Okay.

[00:48:08]Is it visible ma'am?

[00:48:10]>> It's visible Dr. >> Okay.

[00:48:20]First of all, thank you uh Winter Ma'am and Shanka Ser for inviting me for this session. So I am Shanka student uh from SAT. I did my BBS here. So since then I'm happy and privileged to share this uh session uh with sir on stage and uh thank you for everyone else also and welcome to everyone else who are uh in the audience. So uh what we'll be discussing today after the previous session is something completely different that is mostly on the genetic testing part and I'll be dealing today with the cytogenetics and molecular genetics part. Molecular cytogenetics part the proper molecular genetics would be taken in the future session is what uh I understand.

[00:49:09]So before I go uh directly into the different cytogenetics testing, what uh I want to push here is uh we have all heard about this different types of genetic testing, different uh just a second.

[00:49:29]Yeah. So we have uh we often hear about all these different names of genetic test like CMA, Kotype, MLP, QFPCR, SAR, NGS and all. So at uh a lot of times we are not really aware of why all these test exist at all. uh many have uh even when I was doing my MD also I had this misconception of if uh whole genome sequencing can do uh pick up everything why don't we do everything at once and if whole exom uh picks up every coding region then why not do it always why does a sequencing technology not pick up a uh deletion and all those things uh so it is very common uh to have such worries so I think uh a very short introduction on why this matters and why this excess is important. So I'll just briefly go through uh the structure of the nucleus or structure of the DNA. So we know that within each of these cells we have the nucleus and in nucleus uh within the nucleus we have the chromosomes right and within the chromosomes it is the DNA strands which are completely wound again and again into forming these chromosomes right so why this is important is because the testing that we do is entirely depend on uh dependent on what we are actually looking for if it is a problem at a chromosome al level. I'll describe in detail what these are. But if you are looking at a macroscopic or a very large level change, we have a separate set of test. Whereas when we go into deep into the DNA strands if you are looking at the base pair level then the tests are completely different. So different test for chromosomeal level and different test for DNA level. So that's what we'll be describing here. So genetic abnormalities very basically can be divided into chromosomeal level abnormalities and gene level abnormalities. Chromosomeal level abnormalities mean that a huge chunk of the chromosome is either added or it is missing. So here we can see that in down syndrome the whole chromosome 21 is extra. Right? In turn syndrome the whole exom X chromosome is not there. one of the and if you see the dig syndrome this whole region of the 22q11 region is not there. So what happens is that in this huge chunk of chromosome uh there will be hundreds of genes many times. So usually we have two copies of each chromosomes right one we get from the mother and one we get the f get from the father. So two is the normal copy numbers usually but whenever there is a chromosomeal abnormality then the number of copies of these change. So for example in the 22 Q11 region we know the TBX1 gene. Normally everybody will be having two copies of the TBX1 gene but whenever there is a deletion this copy changes into one or if we look into a down syndrome child. Normally everyone has two copies of all these genes in chromosome 21. But in case of a down syndrome child then all these uh genes in chromosome 21 is of three copies. So these are also called copy number variations. Usually we have two copies except in X and Y chromosomes but usually we have two copies but whenever there is a change into one or three or four or zero then the number of copies of GS genes change and that leads to a chromosomal level abnormality and it leads to their diseases. Whereas when we come to gene level abnormalities let's say cell anemia we know that it is because of a point mutation right oralia or cystic fibrosis and all. So what happens there is that the number of copies of the genes is not changing. It is only a sequence change or the point mutation that is happening. So they usually affect only one gene or the activity of a one single gene and it is because of a point mutation. Why this is important is because the test for chromosomeal abnormalities are different and the test for gene level or single gene abnormalities are different. So at chromosomal level we have carotype fisherial MLP and CMA whereas in gene level disease sort to find out sequence abnormalities we have next generation sequencing and SAR sequencing. So today what we'll be discussing is the most important of these chromosomeal level abnormalities and future sessions these would be covered this side would be covered. So today we'll be discussing the test for chromosomal disorders especially cytogenetics and molecular cytogenetics which includes karotype fish and CMA. So what is the difference between cytogenetics and molecular cytogenetics? Cytogenetics means the study of these chromosomeal level abnormalities usually or classically we have always done it in a uh way where we visualize these. So typically kotyping.

[00:54:43]In karotyping what we do we'll just uh do the staining and we visualize these chromosomes under the microscope. So that is cytogenetics. But whenever we do a mo we go into or we get into the molecular level or DNA level either by studying it or targeting it then it becomes a molecular cytoine. For example, in fish it's it's not we do visualize it but we have a probe which goes into the molecular level and binds to the target uh DNA and in CMA we shred the DNA and then read the chromosomeal level abnormality. So fish and CMA usually we call that molecular cytogenetics whereas karotype is the conventional cytogenetics. So before we go into uh each of these tests there are couple of things we should know. So earlier I mentioned that chromosomeal abnormalities usually lead to copy number variations right the number of copies are changed but that is not entirely correct in many of the chromosomeal abnormalities the number of copies might not change for example we very commonly come across something called as balance transllocations or reciprocal transllocations where here we can see that it's chromosome 1 and two one part of the chromosome one has gone to the two and that part has come to the chromosome one. So the net chromatin or the total amount of chromatin is not changing or the number of copies of these genes are not changing. So these also come into chromosomal structural abnormality. Similarly in inversion this is only rotated among itself. So the number of copies of the gene is actually not changing. It is actually a structural abnormality. Why this is important is because uh not all tests can detect these structural abnormalities. So we'll come to which all can detect these. So many tests can detect chromoso uh this uh copy number variations but other tests can uh some test can uh sorry some test can detect structural abnormalities but some cannot detect both of them usually can detect the CNVs based on their resolution.

[00:56:48]So another concept which is important is mosaicism because again we know what mosaicism is. In the early part of embryogenesis one of these cells become abnormal and based on what those cells develop into the same individual is having multiple s uh uh multiple genetic makeup. A few of his cells have a different genetic makeup but uh the rest of it has a different makeup. Again why is this important in this class is because not all tests can detect mosaicism. A few tests can detect it whereas the others cannot. So these two are important concepts before we going go forward. So first we'll directly go into to the most basic test or the uh most commonly and the uh usually prescript test. It is gradually changing but again it is the most widely available chromosomeal test or cytogenetic test currently. So again kerotyping usually we take it in a heperin well in green top and we need a fresh sample because we need to culture the sample. If there are no actively dividing cells the culture phase so we will not be able to perform a proper carotyping. So uh this is the basic of how you can send a sample. So what we do is we culture it and we uh arrest it in the mitosis phase in the mitoic division phase and then we uh fix it on the glass legs. We stain it and just visualize it under the microscope. So it is one of the basic things that we can do. We just uh stain the cells, arrest it. Uh we stain the cells and then we visualize it. So that itself uh uh we understand what can its uh its advantages and disadvantages be.

[00:58:35]As you can see after the staining we see something similar to this in the microscope and we arrange it against a few graphs called idiograms and we'll finally reach to something called as a carioraph. based on the size and type of chromosome we arrange it and we look it and look under the microscope on whether something is abnormal or not. Again it is very obvious that it has its limitation. The main limitation being that we can only detect abnormalities which are large enough to change the banding pattern under a microscope. So that is the main issue that it has.

[00:59:13]Usually it comes to around 5 to 10 mega base. So uh in the monogenic condition or single gene condition we are uh we are checking for one base pair resolution whereas in kyotyping we can only see things which are more than 10 to six base pairs in size. So that is the main issue with kyotyping. It is very easier to do and very widely available. But the problem is with the resolution it is unless the problem is or the chromosomal abnormality is large enough to change the banding pattern we will not be able to detect it. So this the advantage again cheaper it is accessible uh easily accessible and usually very easier to set up also but the lower resolution is the issue and labor intensive and it is highly subjective also because uh uh we need experienced uh people to look at the reporting also because uh uh it takes a lot of practice to identify each of these chromosomes and to arrange them and to find these abnormalities.

[01:00:20]So there are special forms of kotyping.

[01:00:22]now available which can improve the resolution somehow but again not widely available or not uh like uh very uh impressive to overcome its low resolution issue. So when do we do it?

[01:00:35]Whenever we have a non- chromosomal syndrome especially for example if you have a down syndrome child or a typical Edward child or turner syndrome or Kleinfelter and all we can uh do this because again the chromosomes are usually visualized. So any anuploies we can pick it up early. uh this DSDS whenever a baby is there abnormal ambiguous gentilia is there or some internal mullarian abnormality is there and all we want to confirm the sex of a patient or a child we can do a carotyping earlier when CMA was not available it used to be the first line testing for many of these things where chromosomal abnormalities were suspected like multiple organ malfformation facial dysmorphism growth retardation when this kind of a of a uh triad or this kind of a uh syndrome comes when we as a first line test we used to do kotyping but now that the CMA is more easily available many prefer to go for CMA we'll come to that later so again this is a very typical image of a child with Edward syndrome because this overlapping fingers this rocker bottom foot this hyperextended uh hips and all knees we can see and we can see that it is a tisomi 18 child. Again another case where uh so it it always need not be a anuploidity because very typical example is a kredo chart syndrome where it is not an anuploid but the abnormality is large enough that it can be visualized under a microscope. So this is another case of a crudo chart uh baby who was uh suspected to have crudo chart from the uh typical cat like cry and other features and all and in chromosome it has revealed uh uh 5p deletion.

[01:02:32]So another very common scenario that we come across is coupled with the recurrent abortions. So uh again classically for products of conceptions whenever there is a recurrent uh abortions or miscarriages we take the products of conceptions and do a genetic testing or uh so we classically used to do karotyping again uh recently it has been replaced by CMA. Here we can see a case of risomi uh 13 because of a transllocation and in such cases we do the parental carotyping also. Here we can see that one of the parents has this balanced transllocation or uh robotssonian transllocation between 13 and four this 14. This is why the the couple is having recurrent abortion. So again for couple uh PC testing and couples with uh uh recurrent abortions we usually test it uh you you said kotyping especially for parents although for PC testing CMA is an option for parental or couple testing we still have to relay on carotype why because it comes back to the earlier couple of things I mentioned the balanced chromosomeal abnormalities or structural abnormalities this is a structural abnormality This will not be picked up in a chromosomal microarray. Although microarray as we'll see later is a high resolution test. It cannot pick up balanced things. So couples with recurrent pregnancy loss or although in products of conception we can do a CMA for the couple we are expecting something uh asymptomatic or the chromatin to be complete. We do not expect any CNVs there. In such cases we have to go for a carotype only.

[01:04:20]Okay. So this is the uh sort of a summary of what we have uh discussed earlier. If uh uh someone wants they can take a screenshot. Uh so the thing is we do it whenever the change expected is large enough to be seen under a microscope especially for macro deletions and large structural abnormalities etc. Usually it takes around 12 to 14 days.

[01:04:45]Okay.

[01:04:46]So coming to the next uh test that is fish. Fish also is a cytogenetic test but we use a probe.

[01:04:56]So it it works on a simple principle. We need to know the target that we are looking for. So for carotyping uh we did not need to know any target right. We can just visualize every chromosome under the microscope. But whenever we are doing fish, we need to know what target we are looking for. We'll take the probe which is complimentary to that target which binds to the target and produces fluesence. So if based on the number of fluesence we will be able to find out the number of copies. So sorry.

[01:05:31]So that is how it works.

[01:05:34]Uh so fish is also heperent sample. It's a typo there. So but the main thing is that we can send a stored sample especially if there if we are not looking for a metaphase fish. Uh uh if it is for an interface fish we can do an ADA sample also and do a stored sample.

[01:05:52]But again similar to carrier type if we want to do a metaphase fish then we need to have a heperin sample and uh live sample or fresh sample only.

[01:06:04]So how it works is again we denature it.

[01:06:08]We hybridize it with the uh help of fluorescent probe and when then we see it under the microscope. As you can see this is a very typical example of how such a cell would look under the microscope. So it is a the red one is a probe for chromosome 21. So the other one is a control probe. There are two dots only which means that there are only two copies of it. But for chromosome 21 there are three dots which means that there are three copies of it.

[01:06:39]So it's a case of down syndrome. It can also be used to detect fusions sometimes. So this is again not very important for pediatrics but you can know how it is being used. So the red probe is for ABL and green probe is for BCR. So here you can see that there is green plus red plus a yellow fusion in between. So this is the typical Philadelphia chromosome uh or the BCRL gene fusion. Okay. So for oncology for fusions also it can be used otherwise for typically counting the number of copies of your target is what we use it for. The advantage is that it is much more rapid especially when we don't need the culture and it is uh if we know the target the main thing is that if we know the target it is a very good test especially if the probe is available if the probe is not available for a particular region then we can do it but if the probe is available and we know the target then it is a very good test to be done but the limitation is that again only the targeted ones we can do it and some sort of especially in oncology ology and all some sort of subjectivity is there but usually it is a better test. The resolution we usually don't say for targeted test because it's usually a yes or no uh answer. So it is a targeted test. If we know exactly what we are looking for and if the probe is available then it is a good and a quicker test to to run.

[01:08:07]So what are the indications especially for anuploidities for example uh if a baby is born especially we have certain cases where uh uh newborn baby is there and uh hyperlucosetosis is there and we want to see if the baby is having down syndrome or not. Many times it is very obvious in the face but many times it is not. So for rapidly checking for uh tisomi 21 uh and even in other children who are not diagnosed because the treatment of uh tam associated uh leukemias are different from the other leukemas. So there also many times we have to do a rapid unemploy testing for uh uh postnataly sometimes prenatally also when we have a 23 + 4 weaker or 23 plus one weaker mother antiatal we are suspecting some anuploidity we want the report very quickly especially since the inuploidity since the ultrasounds usually if the uh testing we are doing at 23 weeks and all usually we have normal ultrasounds So we have to see for a mosaic also. Are we looking for a mosaic baby uh which does not have any ultrasound abnormalities till around 23 weeks. So we often go for fish testing.

[01:09:27]Uh very common scenarios. Newborn with DSD again we consider uh DSD or disorders of sexual development or ambiguous gen at birth as a social emergency right. So people have to tell their relatives and all uh whether it's a boy or a girl. So we need a quick uh determination of the sex of the baby in case of ambiguous interiors and all we again go for a uh fish testing because stereotyping is going to take up two days. Again uh for other things whenever we know exactly what look we are looking for for the 5pucha test also we can do a fish test if we are if we know specifically that is what we are looking for or digor syndrome or william or anything if we know specifically that is what we are looking for and if we have the probes in house then uh we'll be able to do these tests it's a targeted testing basically so this is the scenario we know what this We have hypogonadism. We have brachidactyl ID is there. Obesity is there. It's a typical case of a parlay syndrome. Here we can see that we have used a probe for PML and SNRPN. The PML probes there are two dots but for SNRPN there is only one dot which means that the SNRPN gene is deleted. It is typically it can be seen in both Pilly and Angelman.

[01:10:54]uh uh uh this based on clinical features we can find that it is a paral release syndrome.

[01:11:01]Next again it's a case with diode syndrome uh to baby corner triangle abnormality hypocalcemia and uh this absent thyimus etc. You can see that there's a red probe is the one to for 22q11. You can see that there is only one dot. This is we can how we can identify this uh uh uh dig syndrome quickly with the help of fish. Again in many other cases where we are thinking just of any chromosomeal abnormality the typical features of chromosomeal abnormality is there but we don't know exactly what chromosomeal abnormality it is. We cannot use it because it is a targeted test. So this is again a kind of a summary for fish testing using fluorescent probes and microscope targeted testing if it's for a well-known uh condition for which probes are easily available or for a new employes and all for prenatal and quick uh reporting etc. we can use a fish test.

[01:12:02]Next is CGH we call it CMA also chromosome and microarray. In that what we are doing is we are shredding the DNA. Until now in both of the cases we were just visualizing the whole thing uh with the probe or without the probe just staining. But in this case we'll be shredding the DNA putting into splitting in very simply put we'll be splitting in into millions of pieces. Each of these pieces will go into a chip or array and there are small wells in each of these uh uh chips and each fragment goes into each of these chips and if the target is there we can say we can kind of think of it like uh fish millions of fish happening parallelly for different targets. It is not like that but you can visualize it like that. And in each of these wells a reaction takes place. And based on how many copies are there within uh uh this uh fluoresence color will change and a higher resolution photograph is taken of all these millions of wells which is turned into final report on whether there is a gain or there is a loss or whether is a neutral number of copies. So this is how microarray works. So the main advantage it it has if you remember karaotyping it was unargeted anything which can be seen under the microscope could be found out irrespective of whether we are expecting something or not. But the main issue was that it was low resolution. So kotyping was a low resolution and uh uh non-targeted testing. But when coming to CMA it is a high resolution unargeted testing. That is why it has replaced carotyping in many of these indications that we commonly see. So even 25 50 or even sometimes five or 10 KB size of micro deletions and duplication can also be picked up in CMA. Advantages that again it is unargeted. We can uh check for multiple areas together. A very high resolution. We can sometimes uh we can usually detect uh LO regions, UPD regions also. But the problem is it is high cost and again like we mentioned earlier low mosaicism or less than 20% mosaicism we earlier p talked about it pitch and carotype since we are pick seeing one cell at a time if we say see 100 cells and 20% is affected or 5% or 10% is affected on an average in 100 cells we'll be able to see that abnormality in around 10 cells right but in CMA unless it's around 20% we tend to miss it. Balance transllocation again we usually miss it because once it is shredded we'll not be able to say whether at what position it is. Let's say a particular portion of the chromosome is shifting from chromosome 1 to chromosome 2. Once shredded it is not traceable back right. So even if there is a balance transllocation or not the report would be normal. So these are the some of the limitations of cment. So indications again uh earlier we mentioned it as the uh we mentioned some indications for kotyping right. So except for uh parental testing in case of recurrent abortions usually in case of pediatric things uh CMA is replacing kotyping as the first line test and whenever there are central abnormalities dysphorism dysmorphism growth failure etc. We saw earlier kerotyping can be used but CMA is a higher resolution test. In addition, it has been the first line test in many cases like unexplained development delay, autism, behavioral issues and uh in case of prenatal ultrasound showing abnormalities and all. It's usually the first line test for detecting chromosomal abnormalities.

[01:16:05]So this is the scenario again have baby having facial dysmorphism, hypotonia, cardiac disease, uh IV etc. And we did the uh CMA for the baby and we found out a 15 Q11 micro division which is the uh typical this uh uh hypotonic baby with prior relay syndrome. Okay.

[01:16:27]Again the prior can be detected in fish also if it's targeted but in this particular cases we were checking for all the chromosome abnormalities because of the uh features but we found it uh in the newborn period itself we found it to be a prior group.

[01:16:44]This was another case again a major issue with uh CMA you can see is that small CNVs can be there even in normal people right? So when whenever we do such a higher resolution array the problem is that many times we'll we'll get this 22q1 micro deletion but we'll get some uh vus similar to in NGS and all we'll get CNVs of uncertain significance also which is usually not problematic but especially in prenatal cases whenever we detect CMVs of uncertain significance it is a difficult dilemma because we usually don't have time to test the parents and all. So again just like with NGS that we will be seeing in the future classes V uses C the CMA having such a higher resolution has this issue of VUSS also. So this is a a summary of CMA it's a non-targeted test it has higher resolution but cannot pick up mosaics and uh this uh balanced diseases balanced abnormalities.

[01:17:51]So this is an overall summary of what I have made in case of chromosomal abnormalities. If you are looking for anuploides, balance transllocations or large CNV, scarotype is good. If it's micro deletions and duplications and it is known that what we are looking for, we can go for fish. It's unknown but usually fitting the spectrum of a chromosomeal disorder, CMA is the better test. Balanced transllocation can be picked up in kyotype and fish. balanced again if it is targeted but uh imbalance can be picked up in all of these mosaic can be picked up only in kotype and fish in CMA anything more than 20% can only be picked up so this is an overall summary of what we discussed uh today so this is again a comparison in case students want to take a screenshot for their exam and all okay so that was today's session I'll just quickly go through uh or 2 minutes of your time to show the lab that we have set up at Kim. So uh we uh the department of medical genetics at Kim was set up two years back in January 2024. It started just with me but now we have two counselors. We have multiple uh we have a rare disease clinic, encogenetics clinic, reprogenetics clinic and all. We uh have we per month we see around 200 patients per uh month.

[01:19:18]It's a relatively small small clinic based on the OPD numbers. Uh but two six seven months back in November uh uh 2025 we took accepted the first sample to our genomics lab. So we have a team a few of the team has changed. It's an old slide but we have a total of around 15 new team members including four PhDs and other technicians working with us on the lab. uh in the last 6 months itself we have crossed uh uh 2,000 samples uh clinical samples all our most of the uh volumes come from uh oncology in addition we are doing reproductive rare diseases also we have all the main technologies available including NGS RTPCR sangar MLPA kotaping and fish and this is the lab it's a total around 3,000 square ft area so most of the uh equipments we have. You can see our staff working with the cytogenetics part and we have the molecular lab and we have a dedicated bionformatics team also. We have a bioninformatician and two genome analysts also. We recently crossed the 2,000 sample mark and uh we are uh planning to expand further because the number of volumes are also uh increasing very steadily. So uh what I wanted to uh say was that we now have such a uh fully fledged lab in trander itself. So if you want to collaborate in some way we'll be happy to collaborate. And one more thing is that we also have launched uh a clinical genetics platform because uh the accessibility of clinical genetics has been a major issue across uh India. So we have a team of 10 people. It is also expanding mostly we'll have around 17 clinical jists by the end of next month across India mostly will be available via online consultation and also we have pan India sample pickup and testing and all we'll help you with all your clinical queries help you uh uh choose the test we'll help the test uh to get the test done we'll handle the post test counseling we'll consult with the patient and also we'll talk directly with you on what are the next steps to be taken on managing the clinical uh patient also all uh as a remote team or mostly all of our services are online with the help of all these geneticistes.

[01:21:56]So again hoping to collaborate through that also thank you for patient listening.

[01:22:08]Okay, thank you. Thank you Rashan for the insightful talk on uh cytogenetics and molecular cytogenetics.

[01:22:17]Now we have um uh minita any questions in the chat box? Sir, >> there is one question sir. How I think it's to madam. How to interpret as carnitine profile in a TMS report?

[01:22:35]Which genetic testing is ideal and cost effective to diagnose?

[01:22:42]>> Very interesting question. If I have to teach you how to interpret as caretian profile, I think tanker sir has to you know give me one more opportunity to go through on interpretation of TMS. Um however uh I can kind of uh say that in the TMS you get two two kind of profile.

[01:23:02]One is the amino acid profile another one is the asel cardian profile. Amino acid profile per se would help us to diagnose ura cycle disorder and amino acidopathies. Whereas asel cararnitine profile per helps us to diagnose organic acidas and fatty acid oxidation defect.

[01:23:21]We remember by O and O. So odd chains will be elevated in the asel carnetin itself. There will be odd chains as well as even chains. Asil is the carbon group and carnetin is the one which gets attached to the asel group. So whenever we get an elevation of octane asyl carnitine then that is suggestive of organic academia like C3 elevation happens in propionic and methyl malanic C5 in isoalic C5 DC in glutaric etc. uh whereas uh EF even chain elevation such as fatty acid oxidation defect depending on which uh you know chain is elevated we can come to a diagnosis whether it is a short chain acco deficiency or medium chain or very long chain um or whether there is a transport defect. So if you ask me one biomarker assets C4 is for short chain, C8 is for MCAT, C14 and C14 is to1 is for real CAD and you know we also calculate some ratios uh to kind of make uh it kind of clear and interested. So E F so even change is for fatty acid oxidation defect. Regarding the genetics um you know as such we uh the previously we have done see the for example the propionic academia which was one of our first few children whom we diagnosed maybe in 201 12 or 2013. So that time we used to send it to Germany for a single gene like you know propionic academia uh gene alone we sent uh but that is not going to be cost effective because propionic academia per se is caused by two genes PCCA and PCCA B. So uh you know if you have to do two things then it turns out to be costly. So majority of the time we we utilize the whole exom sequencing. Uh if there is a screen positive we do newborn screening for all the newborns born here in our hospital.

[01:25:38]All live births undergo uh the extended TMS testing as well as biotinis and galactomia along with thyroid and uh you know cah for them. We do have uh what is known as newborn screening gene panel to make sure that all the genes which all the disorders which are detected by screening or 100% cover coverage happens because if you ask like whole uh exom sequencing some of the uh screen detectable genes may not show 100% coverage to make sure that we have 100% coverage for all the genes which are detectable by newborn neon screening. Uh we do what is known as newborn uh screening gene panel.

[01:26:29]So by and large the answer to your question is we utilize whole exom sequencing.

[01:26:39]>> I think that is clear. There's one more question ma'am. Which lab is good for doing TMS and GCMS?

[01:26:47]I think see more than um lab you should be able to you know interpret you need some help in interpretation um see there could be I have seen reports where depending on whatever metabolite is they don't kind of you know think or put things together it may not be interpreted by a metabolic consultant or metabolic specialist what happens is If leucen is elevated they'll say leucen is elevated it is msud c8 is elevated it is mcat. So everything will be there they want you know try to put things together. Um say for example there is glutaric acidia type 2 or m deficiency multiple as uh you know coa dehydrogenous deficiency where you can get glucin elevated as well as short chain medium chain and longchain elevated all can get elevated but you need that kind of expertise to uh interpret the report. So I would recommend you know in the lab where you can have a metabolic specialist or metabolic expert interpret your report will be better and you as a clinician should gather little bit knowledge on that to be complimentary to each other.

[01:28:12]So if you are asking me where we are sending we in Navi Mumbai run by Dr. Okay ma'am. Next question is what changes we can see in LCH from TMS?

[01:28:30]Okay. As I said again there will be C16 O elevation uh and C18. So very long chain hydroxy because it is L chad right? It is not LCAD it is L chad. So C16 O and C18 O will be elevated. U by and large we won't be able to differentiate by biochemical testing LCHAD or trifunctional protein deficiency.

[01:29:00]Molecular testing would help to differentiate these two.

[01:29:10]Hope I have answer Dr. Sumea.

[01:29:14]If that's I think that's clear.

[01:29:18]Any more questions?

[01:29:20]Those you who can also unmute and ask.

[01:29:34]>> Thank you Shinas.

[01:29:41]Anything to add?

[01:29:43]>> Nothing more to add. I was wondering how much extensive is the knowledge of both the speakers and many of the things which were covered was not known to me.

[01:29:54]Excellent talks Dr. What about the perfectly normal children, late onset problems, episodic attacks here, episodic phenomenon where we'll be considering mitochondrial disorders in many of the episodes triggered by the stresses and all. Uh so how long we should be suspecting this thing? How long are going to supplement the the the main principles? said few of them are imminently treatable by supplementing the vitamins or co-actors that's what we are doing. So how long this can happen?

[01:30:33]Uh adolesence is there any limit?

[01:30:39]>> By and large mitochondrial disorders if you take a blanket of mitochondrial disorders mitochondrial disorders can be of nuclear inheritance or mitochondrial inheritance per se. Uh the majority of them like we see in newborn or in like younger infant or of nuclear inheritance. They are actually autotosomal recessive which which is very severe and they present in the neonatal period itself. Whereas a mitochondrial disorder due to mitochondrial inheritance they can present even up to in adult or adolescent as such. Now the the clue here will be multi-organ involvement as such. So you you will be unexplained and multiorgan dysfunction.

[01:31:22]So this is one aspect of it. When we are talking about movement disorder, it is a very interesting perspective. So we do kind of uh you know put the movement disorder as chronic. Baby would have received some phototherapy for some daughters of 15 or 16 because like we are not able to explain the distonic moment etc. We might kind of put that as a diagnosis. Whereas actually there could be neurotransmitter disorders or terine defects. um many of the metabolic disorders can present with abnormal movements or donia.

[01:32:00]I can share the paper which I have uh uh I have written a review article in Indian journal of tactical pediatrics where there is a wide range of disorders which can present as CP mimics cerebral paly mimic becomes known case of cerebral paly. No one would have explored into whether there was perinatal asexia, whether there is a baby who was born prematurable to a you know the brain dam the neonatal period without knowing that you would put a cerebral paly and then it becomes a cerebral paly. I do have seen baby coming as diplesia diagnosed to have arginemia because my interest is in small molecule disease. I restrict my I am basically I'm a neonologist but I restrict my you know thing to small molecule disease it's a diet based treatment that is my area of interest but I do see because of reference some lysosomal storage peroxal disorders those also I do see um so basically we have to categorize in our mind whether they are treatable or not not treatable so we have to all the opportunity we have to look forward for something treatable.

[01:33:20]>> What is the role of you said metronol in OD? How it's working?

[01:33:25]>> Sir um um the we have to our one of the primary principle is to reduce the substrate.

[01:33:33]Um so when we give metronidazol what happens is there will be that bacteria which produces ochain fatty acids. As I said in the cell carine profile also usually our fatty acids whatever is there in the human or even chain fatty acids like C8, C10, C12, C14 etc. Whereas the gut bacteria can produce odd chain fatty acids like C7 which which further breaks down to C3 and C4. C3 is nothing but propionic acid. So we are pl we are actually decreasing the substate which is the propionic acids um by providing metronidolur but if you are going to give metronidosurol through one month then there are like lot of side effects associated with it. So we cycle it. We give one week and give three weeks of drug holidays. Then again so we tell the parents to remember as calendar month 1 to 7 you give metronome for all the other days you give drug holidays.

[01:34:38]That is how we counel the parents. They also understand. So the reason behind it is it reduces the off chain fatty acids which in turn is produced by the bacteria.

[01:34:52]>> Thank you madam do Dr. Roshan what about the uh my question is to the roshen Dr. Corrosion is there.

[01:35:02]>> Uh yes sir.

[01:35:03]>> Corrosion MLP and PCR was not covered.

[01:35:06]Uh or is it a >> covering later?

[01:35:09]>> Yeah. Uh I think it will be covered in the later session.

[01:35:15]>> Other session second session part two.

[01:35:17]So that will be covered.

[01:35:19]>> Molecular diagnosis will be cover MLP sequencing other.

[01:35:23]>> Okay. Okay. Excellent. Excellent Roshan.

[01:35:25]You made it very simple understandable for even the people like Thank you sir.

[01:35:30]Sure. Excellent talk s >> there's one question for Dr. Roshan. Any genetic lab in South India during luning genetic testing like exom sequencing I think yeah at reasonable cost and reliable results it's a different thing sequencing.

[01:35:50]>> So uh again the quality I think has to be checked for each of these labs separately. For example, in our center, we are charging around 18,000 for exom sequencing because we are in control of the quality. We know what happens to the sample. Again, there are lots of uh uh ways to cut the cost of uh these test also. If you are supposed to run 50 samples together, if you run 100 samples together also, we'll get the data. But the data would be of subpar quality. We can use diluted reagents also, half dilution. So again lots of uh ways are there to cut the cost of exumes. Again at the same time there are large machines that are currently available now which also can do it at a lower cost. So whenever someone comes to you with a lower cost whether they are using the cost cutting measure or whether they are using actually a good machine or a high-end machine it is not very easy to find out. We have seen the most popular labs also having gaps especially when there are very typical uh cases and we don't get a report. We take the data and see we'll see huge gaps in the data especially in our target region. We have had cases where we send two samples and the reports were opposite to each other.

[01:37:12]The first person got the report of the second person and second got the report of the first person also. And initially we thought it might have been a mistake from our side but the lab itself cleared that it has been a confusion from their side. So uh what what all things can be under our control and uh how much of a cost cutting they are ethically able to do is of a question always uh it is better that all the uh testing being done under your own lab or inhouse lab you have full control over it. Again uh it is difficult to tell any single lab uh all these labs I'm not a fan of any of these labs because most of these labs we have had bad uh uh uh experiences with but again for the time being what we can do is that until the decentralization happens and the uh if the just like pathology was earlier centralized and now it is there in all hospitals in future mostly genetics will also come to most of these hospitals.

[01:38:18]Until then we can use most of the labs which are giving at a very affordable rate. But any of these are better than each other. It is difficult to say all these labs use these cost cutting measures. So uh right now the choices based on the ease and the cost only.

[01:38:35]Anger sir >> can I add a point? Yeah. Um so am I audible?

[01:38:45]>> Yes. Yes. Yes. Um yes actually uh you know what is what is the cost which we are paying for to terminate the diagnostic ordy which the parents would have kind of undergone like going from pillar to post not knowing what it is. I think that is the cost they pay for. So rather than we we should uh we should not take uh you know in that aspect and as far as M is concerned we we always tell that you know you prove phenotypically and then go for molecular do not jump for molecular always history physical examination biochemical testing followed by molecular. So if your phenotype pick uh birth each and every bird what you write is very important to pick up things. uh just I wanted to add on regarding the cost our fight with us for cost. So we explain them that you know uh many of our prenatal sample why this much cost why this cost etc. So we have to explain our physician also uh in a very similar fashion. So instead of like not knowing let them know what is the diagnosis especially if they are terminating >> you said routinely you are doing the unital genetic thing how much is the cost involved and can be adopted especially those who can afford in Kerala that's why >> definitely yes sir Kerala IMR is so less it is time for Shanka sir will agree with me it is time for us to move into preventive aspect Who says that when you have a reasonably lower uh infant mortality, it is time for us to think about genetic preventive aspects etc. We charge around uh sir we have two units. One is the corporate side and uh medical college site.

[01:40:39]Medical college site is a free site we call like no big charges etc. uh for medical college side it will be around 1,900 for the TMS along with biotin and galactosemia for the corporate side it will be around 28 2,800 but less than 3k so it is less than 2k for medical college side and less than 3k for um medical center um so whether we can adapt yes we can adapt uh now uh We do have you know lot of facility in terms of medical food is available. Um and we do have diagnostic evaluation available. Um and even the parents they they kind of uh uh they they plan for a very small family. So they want everything to be done for the kids. I would say so many uh you know private um hospitals they do photo shoots etc. It will cost only like less than those photo shoots which are done. I always tell for naming ceremony anyone will spend so much instead they can spend for newborn screening so that the life of the newborn is very very much for biotin it is deficiency it is no cost at all just biotin tablet that's all they need but it it saves life it prevents neonal seesures uh it there is no developmental delay etc. So it it makes a very huge difference to the family and the newborn per se. So I would say that Kerala should go for it and all the starboards here should take it forward.

[01:42:22]>> I'm happy to help in case if there is anything I need to do I'm happy to help.

[01:42:27]Sir >> especially that perodoxile phosphate I I I was not aware of the injections available here. Because and the oral preparations and all the biotin is something very very easily doable and the other thing also oral preparations if it available is it available in Kerala also sa >> yes sir yes >> I didn't use it okay >> it's not freely available little bit difficult but it's available >> s even the re nerve where there is B12 paradoxin is also there we just need to see there so re nerve plus neurocrine gold like that normal neurocrine contains only B12 whereas neurocine gold there is some additional birds but right now they are kind of cutting down on combination medications a lot so I do not know what will happen in future but having said that at least this point it helps if you do not have an IV preparation you do not have to worry sir either the benadon tablet or the punk tablet give it through og tube that's all is needed So the same IV preparation you give through OG tube only thing is the EG when we connect bedside it will not have an immediate response but the baby will respond subsequently one have to remember that any seizures even paradox independent seizure can be partly controlled by mizol so that does not mean that the baby does not have paradox dependent seizure little bit considering treatable causes of seizures is very important In that case you use a 200 migression or 40 mig benadon belong or venadon >> sir belong is 100 mg benadon is 40 mg even 40 if you do not have belong benadon itself two and a half tablets you can give we give 100 mig three times sorry 100 mg 3 days continuously irrespective of the response in our unique protocol is 100 migram day 1 100 migram D2 day 2 and 100 migram day three before we uh kind of say that there is no response at all.

[01:44:36]>> There also you get the EG response. You test the EG response before and after oral preparation.

[01:44:42]>> Sir, we have continuous EEG monitoring.

[01:44:44]So all our newborns who have seizures, they will be connected to the EG monitor bed site.

[01:44:53]>> Thank you both of you.

[01:45:02]is banned down total 100 mg in a day for first three days following which you can give around 15 to 25 mg per kg of pyrooxin. So if it is a 3 kg baby it will be around you know for 40 mig tablet one and a half something like that.

[01:45:32]Thank you ma'am.

[01:45:35]I think uh there are no other doubts.

[01:45:38]So we come to the end of the session. On behalf of Perala Genics chapter I sincerely acknowledge the efforts of office buyers president Dr. Gandhumas sir secretary Dr. Bobby Moen and treasurer Dr. for all the help to make this happen and my heartfelt thanks to those south vice president for joining us and my sincere gratitude to the guest speaker and the expert in metabolic medicine Dr. Mama Mahishi madam for accepting our invitation and we appreciate your dedication and efforts you put to the success of this session.

[01:46:13]Thank you madam for the elaborate and uh useful presentation. Next I thank the our next speaker Dr. Ocean Daniel the dynamic one for the excellent presentation and sharing the valuable points in the lab interpreted laboratory interpretation of psychoggenetic and molecular cytogenetic test. I would like to thank Dr. Shandas sir for the expert in genetics for the constant support and guidance toward this program and I thank Dr. Gin sir chairperson genetics chapter for for the contribution to the success of this program. May I also thank all the senior faculties sir pushutin sir parvati ma'am uh for the your valued presence and I thank all the participants and have a good night all of you >> just one suggestion before winding up the experts in these rare disorders and gent groups once in a while only get having extensive experience in this selected area and I would like to have the em the email in one of the slides maybe the last slide or the beginning slide experts email maybe because we can directly interact later Dr. Omar Dr. Roshan anybody in the next future presentation also one slide for with the >> very good suggestion very good suggestion >> thank you good night >> it's not sir >> all of you thank you so much >> thank you >> thank you sir >> thank you good night >> thank you shanker >> thank you everyone good night >> thank you all good night >> thank you ma'am Um thanks.

[01:48:00]>> Thank you Shanka sir for invading me.

[01:48:03]Thank you Gir siras.

[01:48:07]Thank you