Rametrix (R) molecular urinalysis system

Urinary tract disease (UTD) is a major global health problem, affecting affluent and disadvantaged countries and regions alike through communicable and non-communicable etiologies.  Chronic kidney disease (CKD), affects more than 10% of the global population (Ly, et. al., 2019) and 15% of the US population (CDC, 2021), with urinary tract infection (UTI) registering more than 150 million cases (worldwide) seen annually with the prevalence of UTI likely many-fold higher than this figure, since many infections go undiagnosed (Ozturk, et. al., 2020; Zeng, et. al., 2022).

We are addressing the challenge of detecting and managing urinary tract diseases (UTDs) in Sub-Saharan Africa.

Acute and chronic UTD is a major health problem in Africa/Sub-Saharan Africa, with many remaining undetected and without treatment, leading to progression to late-stage chronic diseases that are difficult to treat - with greater economic cost on the health system for clinical management. Consequently, undetected and untreated UTD leads to suffering, diminished quality of life, premature death, and substantial economic loss in many low- and middle-income countries (Stanifer, et. al., 2014; Kayange, et. al., 2014; Glaser, et. al, 2015; Abd El Hafeez, et. al., 2018; Hodel, et. al., 2018; George, et.al., 2019).

Major contributors and risks for development and progression of urinary tract disease in Sub Saharan Africa include prevalent, serious infectious diseases (schistosomiasis, human immunodeficiency virus disease [HIV], malaria, among others), antibiotic-resistant bacteria, poor water quality, acute and chronic malnutrition, environmental contamination of water/food/air, and the effects of urbanization and lifestyle (diabetes, hypertension). UTDs have particular consequences for poor maternal health and adverse perinatal outcomes in developing countries with skewed high rates of hypertensive disorders and gestational diabetes in pregnancy which are difficult to screen and treat with consequent poor pregnancy outcomes. (Gill, et. al., 2009; Peer, et. al., 2014; Glaser, et. al, 2016; Da Silva, et. al. 2017; Ekrikpo, et. al., 2018; George, et. al., 2019; Hamilton, et. al., 2020; Aula, et. al., 2021;) 

A lack of access to healthcare is a major risk factor for disease development and progression. Prompt and accurate detection of UTD in low- and middle-resource environments is problematic - but is a key to successful early disease detection and early institution of clinical management. Current methods of detection include laboratory sampling (blood, urine) and imaging of symptomatic patients, but these methods of detection require relatively sophisticated facilities and can be costly. Such methods of detection are not scalable for population screening, and routine personal health monitoring/healthcare staff may not be readily available for disease prevention and monitoring in the general population.

We will implement inexpensive, rapid, and remotely deployable detection of UTDs.  This will inform actionable follow-up, treatment, and overall disease/health management.

References:

Abd El Hafeez S, Bolignano D, D'Arrigo G, et al. “Prevalence and burden of chronic kidney disease among the general population and high-risk groups in Africa: a systematic review,”

BMJ Open 8:e015069. 2018 doi:10.1136/bmjopen-2016-015069

Aula, OP, McManus, DP, Jones, MK, Gordon, CA, :Schistosomiasis with a focus on

Africa,” Trop. Med. Infect. Dis. 6,109. 2021 https://doi-org.ezproxyberklee.flo.org/10.3390/

tropicalmed6030109

CDC – Centers for Disease Control and Prevention, 2021; https://www.cdc.gov/kidneydisease/publications-resources/CKD-national-facts.html

Da Silva, GB, Pinot, JR, Barros, EJG, Farias, GMN, Daher, ED, “Kidney involvement in malaria: an update,” Rev Inst Med Trop São Paulo 59:e53, 2017 

Ekrikpo UE, Kengne AP, Bello AK, Effa EE, Noubiap JJ, Salako BL, et al., “Chronic kidney disease in the global adult HIV-infected population: A systematic review and meta-analysis,” PLoS ONE 13(4): e0195443. 2018  https://doi-org.ezproxyberklee.flo.org/10.1371/journal. pone.0195443 

George, JA, Brandenburg, J-T, Fabian, J, Crowther, NJ, Agongo, G, … H3 African Consortium,  et. al., “Kidney damage and associated risk factors in rural and urban sub-Saharan Africa (AWI-Gen): a cross-sectional population study,” Lancet Glob Health.December ; 7(12): e1632–e1643. 2019 doi:10.1016/S2214-109X(19)30443-7 

Gill, GV, Mbanya, JC., Ramaiya, K.L. et al. A sub-Saharan African perspective of diabetes. Diabetologia 52, 8–16. 2009. https://doi-org.ezproxyberklee.flo.org/10.1007/s00125...

Glaser, N, Deckert, A, Phiri, S, Rothenbacher, D, Neuhann F, “Comparison of various

equations for estimating GFR in Malawi: How to determine renal function in resource limited settings?,” PLoS ONE 10(6): e0130453. 2015. doi:10.1371/journal.pone.0130453 

Glaser, N, Phiri, S, Bruckner,T, Nsona, D Tweya, H, Ahrenshop, N, Nuehann, F, “The prevalence of renal impairment in individuals seeking HIV testing in Urban Malawi,” BMC Nephrology 17:186. 2016 DOI 10.1186/s12882-016-0403-7 

Hamilton, SA, Nakanga, WP, Prynn, JE, Crampin, AC, Fecht, D, et. al., “Prevalence and risk factors for chronic kidney disease of unknown cause in Malawi: a cross-sectional analysis in a rural and urban population,”  BMC Nephrology 21:387. 2020 https://doi-org.ezproxyberklee.flo.org/10.1186/s12882-020-02034-x 

Hodel NC, Hamad A, Praehauser C, Mwangoka G, Kasella IM, Reither K, et al., “The epidemiology of chronic kidney disease and the association with non-communicable and communicable disorders in a population of sub-Saharan Africa,” PLoS ONE 13(10): e0205326. 2018 https://doi-org.ezproxyberklee.flo.org/10.1371/journal.pone.0205326

Kayange, NM, Smart, LR, Tallman, JE, Chu, EY, Fitzgerald, DW, Pain, KJ, Peck, RN, “Kidney disease among children in sub-Saharan Africa: a systematic review,” Pediatr Res. 77(2): 272–281. 2015.doi:10.1038/pr.2014.189. 

Lv, JC, Zhang, LX, “Prevalence and disease burden of chronic kidney disease,”  Adv Exp Med Biol. 1165:3-15. 2019 doi: 10.1007/978-981-13-8871-2_1. PMID: 31399958)

Öztürk, R, Murt, A,” Epidemiology of urological infections: a global burden,” World J Urol. 38(11):2669-2679. 2020 doi: 10.1007/s00345-019-03071-4. Epub 2020 Jan 10. PMID: 31925549. 

Peer, N, Kengne,A-P,  Motala, AA, Mbanya, JC, “Diabetes in the Africa region: An update,” Diabetes Research and Clinical Practice, 103 (2): 197-205, 2014 ISSN 0168-8227,https://doi-org.ezproxyberklee.flo.org/10.1016/j.diabres.2013.11.006.

Stanifer, JW, Jing, B, Tolan, T, Helmke, N, Mukerjee, R, Naicker, S, Patel, U, “The epidemiology of chronic kidney disease in sub-Saharan Africa: a systematic review and meta-analysis," Lancet Glob Health 2: e174–181. 2014 

Zeng, Z., Zhan, J., Zhang, K. et al., “Global, regional, and national burden of urinary tract infections from 1990 to 2019: an analysis of the global burden of disease study 2019,” World J Urol 40, 755–763. 2022.

We have developed and tested a simple, semiautomated technology for the evaluation of kidney function and detection of UTDs. The technology is based on Raman spectroscopic evaluation of urine samples and is termed Raman molecular chemometric urinalysis (Rametrix®). 

Raman spectroscopy is a method for determining the molecular composition of liquids (such as urine) and solid materials (powder mixtures, for example).  Irradiation of the specimen (urine, for example) with laser energy, deforms/stretches/bends/vibrates the chemical bonds of thousands of different molecular components in the specimen.  As soon as the laser irradiation is stopped, these chemical bonds return to their resting state, while releasing small amounts of bond deformation stored energy.  This energy release (photons) can be detected at different wavenumbers and can be used to create a Raman spectrum that represents the ‘molecular fingerprint’ of the liquid.  Comparing one Raman spectrum from a healthy person, for example, with the Raman spectrum from a patient with suspected disease, allows detection of differences in ‘molecular fingerprints’ that characterize diseases (see below). Raman analyzers have become inexpensive (~$5-15K USD) over the past decade, and hand-held models now exist, allowing deployable applications.  We will use this attribute to bring accessible and affordable UTD detection (and treatment options) to populations in sub-Saharan Africa.

Our Rametrix® technology is proof-of-concept validated and published in scientific journals.  It consists of three components: an analytical device containing sample holders and a Raman spectrometer, computational algorithms to process raw Raman spectra to useful information, and a series of reference databases that contain a library of Raman spectra from historical samples.  The reference database spectra are the product of analysis of thousands of urine specimens from healthy volunteers and from patients with diseases that affect the urinary tract specifically and the body, as a whole.  It is very important to understand that new reference databases/data libraries are built “on site” when the samples are analyzed – allowing comparison of samples/spectra from local populations.  These databases and data libraries grow and grow, as more specimens are added, eventually expected to contain the results of analysis of hundreds of thousands of specimens, representing the regional population and temporal conditions.  Data interpretation is not reliant on historical datasets, collected from different populations (North America and Europe, for example), that have been historically biased against datasets from low- and middle-resource countries.

The process of analysis is very simple and illustrated below!  Patients urinate into a cup and some of the urine (1-5 ml) is then transferred into a reusable glass tube that is inserted into a holder in the analyzer.  The analyzer lid is closed and the laser is then activated (up to 15 times) and the raw Raman spectrum produced electronically, by detecting of the Raman scatter radiation.  This electronic spectrum is then computationally normalized (to eliminate artifacts) and these result are sent electronically for comparison with other reference Raman spectra in the database/data library.  Artificial intelligence algorithms are used to find particularly prominent and important differences between a single patient specimen and the reference databases/data library, as well as searching for trends and differences between many types of samples (male vs female, young vs old, healthy vs diseased, acute disease vs chronic disease, and so forth).

Figure 1.  Overview of analysis of patient urine by Raman spectroscopy.

 A single patient urine sample (5ml) can be screened for the presence of UTD, UTI, acute kidney injury, connective tissue disease, diabetic kidney disease, CKD, bladder inflammation, and urinary tract cancer (see References, following this text section). The analysis costs pennies per measurement (consumables) and results are available within 15-30 minutes (done, as previously stated, by electronic comparison of patient spectral results with reference database of >2000 spectra). The technology can quantify glomerular filtration rate (GFR), proteinuria, and hematuria in a single sample.  It has been extensively validated (see References cited below) 

We propose development and deployment of our Rametrix® technology (portable, single- and multi- sample custom-built analyzers equipped with laptop computers, software, database applications) in Malawi - for improving detection and management of UTDs. This is technology that can be used for single-patient analysis or scaled up for mass (routine) screening of urine samples. A version we developed for the automated analysis of potentially thousands of urine samples per day is shown in Figure 2 below.  The simple analytical devices for collection of Raman spectra are rugged, tolerant of a variety of environments (hospitals, distributed regional clinics), operate at ambient temperatures, and do not require highly trained personal to operate and maintain them. The systems can be operated on solar/battery power, if needed. 

Figure 2.  The Rametrix® AutoScanner (left, middle) for automated analysis of thousands of urine samples.  Commercial small footprint scanner (right) for analysis of single urine samples for UTDs.

Development of a Team of dedicated personnel (physicians, healthcare professionals, engineers, students) will make the adoption of this technology a reality. Our Solution Design Team is in place in the US (R&D, manufacturing) and the Solution Delivery Team is in place in the US and Malawi (clinical application of the technology for healthcare - this is more fully described in“How are you and your team well-positioned to deliver this solution?.

References: 

Carswell, W, Guruli, G, Tracy, A, Xu, Y, Du, P, Senger, R, Robertson, J, “Raman Spectroscopy as a non-cytological detection and quantification urinalysis method for microhematuria in human urine,” Applied Spectroscopy, Feb 2022, On-line, DOI: 10.1177/00037028211060853 

Fisher, AK, Carswell, WF, Athamaneh, AIM, Sullivan, MC, Robertson, JL, Bevan, DR, Senger, RS, “The RametrixTM LITE Toolbox v1.0 for MATLAB®,” J Raman Spectro  49 (5): 885-896, 2018

Huttanus, H, Vu, T, Guruli, G, Tracey, A, Carswell, W, Said, N, Du, P, Parkinson, B, Orlando, G, Robertson, J, Senger, R, “Raman Chemometric Urinalysis (Rametrix™) as a screen for bladder cancer,” PLoS One. 2020; 15(8): e0237070. Published online 2020 Aug 21. doi: 10.1371/journal.pone.023707016.

Kavuru, V, Vu, T, Karageorge, L, Choudhury, D, Senger, R, Robertson, J, “Dipstick analysis of urine chemistry: benefits and limitations of dry chemistry-based assays,” Postgrad Med Published on-line Oct 19 2019 https://doi-org.ezproxyberklee.flo.org/10.1080/00325481.2019.1679540

Robertson JL, Senger RS, Talty J, Du P, Sayed-Issa A, Avellar ML, et al. “Alterations in the molecular composition of COVID-19 patient urine, detected using Raman spectroscopic/computational analysis,” PLoS ONE, July 2022 17(7): e0270914. https://doi-org.ezproxyberklee.flo.org/10.1371/journal.pone.0270914

Senger, R, Kavuru, V, Sullivan, M, Gouldin, A, Lundgren, S, Merrifield, K, Steen, C, Baker, E, Vu, T, Agnor, B, Martinez, G, Coogan, H, Carswell, W, Karageorge, L, Dev, D, Du, P, Sklar, A, Pirkle, J, Orlando, G, Lianos, E, Robertson, JL, “Spectral characteristics of urine specimens from healthy human volunteers analyzed using Raman Chemometric Urinalysis (Rametrix™),” PLoS One, Published: September 27, 2019 https://doi-org.ezproxyberklee.flo.org/10.1371/journa... 

Senger, R, Robertson, J, “The Rametrix PRO ™ Toolbox V1.0 for MATLAB®,” Peer J, 3:35799:2:0, 2019 https://peerj.com/articles/8179

Senger, R, Sullivan, M, Gouldin, A, Lundgren, S, Merrifield, K, Steen, C, Baker, E, Vu, T, Agnor, B, Martinez, G, Coogan, H, Carswell, W, Kavuru, V, Karageorge, L, Dev, D, Du, P, Sklar, A, Pirkle, J, Gulich, S, Lianos, E, Orlando, G, Robertson, JL, “Spectral characteristics of urine from patients with end-stage kidney disease, analyzed using Raman Chemometric Urinalysis (Rametrix™),” PLoS One, Published: January 10, 2020; pone.0227281

Senger, RS, Scherr, D. “Resolving complex phenotypes with Raman spectroscopy and chemometrics.” Curr. Opin. Biotechnol. 66: 277-282, 2020 doi:10.1016/j.copbio.2020.09.007

Senger, R, Sayed Issa, A, Agnor, B, Talty, J, Hollis, A, Robertson, JL, “Disease-associated multimolecular signature in the urine of patients with Lyme disease, detected using Raman spectroscopy and chemometrics (Rametrix™)” Applied Spectroscopy, Feb 2022, On-line, DOI: 10.1177/00037028211061769

Senger, R, Sayed Issa, A, Agnor, B, Talty, J, Hollis, A, Robertson, JL, “Disease-associated multimolecular signature in the urine of patients with Lyme disease, detected using Raman spectroscopy and chemometrics (Rametrix™)” Applied Spectroscopy 76 (3): 284-299, 2022 DOI: 10.1177/00037028211061769

Xu Y, Du P, Senger R, Robertson J, Pirkle JL. ISREA: An efficient peak-preserving baseline correction algorithm for Raman spectra. Applied Spectroscopy. 2021;75(1):34-45, 2021 doi:10.1177/0003702820955245

Xu, Y, Du, P, Senger, R, Robertson, J, “Sparse logistic regression on functional data,” Stat Interface 15(2): 171-179, 2022 

Zu, TNKK, Athamneh, AIM, Collakova, E, Robertson, J, Hawken, T, Aardema, C, Senger, RS, “Assessment of ex vivo perfused liver health by Raman spectroscopy.” Journal of Raman Spectroscopy. 46:551-8, 2015

In preparation:

Huttanus, H, Martinez, G, Coogan, H, Zhang, S, Donaghy, T, Du, P, Karageorge, L, Robertson, J, Senger, R, "The effects of sterile-filtration on urine molecular composition and degradation when analyzed by Raman Chemometric Urinalysis (Rametrix™)" (in preparation)

Kavuru, V, Senger, R, Robertson, J, Bhagwat, M, McNeil, L, Wines, E, Dev, D, “The molecular composition of urine, detected with Raman spectroscopy and chemometric analysis, can be used to differentiate clinical presentation and renal pathologies in patients with diabetes mellitus” (submitted, Peer J, July, 2022)

Scherr, DM, Guruli, G, Robertson, JL, Senger RS, “Screening urine for chronic kidney disease and bladder cancer with a quinone-luminol assay,” (in preparation, J Appl Spec)

Senger, RS, Du, P, De la Torre Campos, D, Carswell, W, Webster, K, Sullivan, M, Gong, J, Robertson, J, “Assessment of urine specimen storage conditions using Raman Chemometric Urinalysis™ (Rametrix™),” (in preparation, J Appl Spec)

Thyolo District in Southern Malawi is Malawi’s 5th most densely populated district with a population of about 721,000 people (Gov Malawi, 2022a). The population is rural consisting largely of subsistence farmers and farmworkers of commercial farms. Thyolo District Hospital (TDH) is one of two secondary health facilities in the district and the only government one (the other is a CHAM facility). In addition to the large catchment within the district TDH also services communities from the adjacent Chiradzulu and Mulanje districts. The nearest tertiary/referral hospital for the region is the Queen Elizabeth Central Hospital in Blantyre, over 40km away. Although tertiary/referral facilities are supposed to provide specialist health services at regional level and also provide referral services to district hospitals within their region, in practice, however, around 70% of the services they provide are either primary or secondary services due to lack of a gate-keeping system (Gov Malawi, 2022b). The high patient burdens at referral hospitals and transportation costs associated with referral mean that most often only the most critical cases or those that are deemed to be in need of specialized care are referred. As a consequence, many referrals are made too late and by the time patients arrived for consultation at the central hospitals (Kamuzu Central & Queen Elizabeth Central Hospitals) their communicable and non-communicable diseases were often so advanced they were not treatable.

The provision of specialized diagnostic capacity in a secondary facility is critical as these facilities play an important role in receiving patients from the community and primary levels of care where a majority of the population will first present for care. The significance of this is heightened in light of the limited capacity of these facilities to provide laboratory diagnostic services resulting in routine misdiagnosis of even the most important diseases such as febrile illness  (Peterson, et. al., 2021). There is an increasing burden of non-communicable diseases (NCDs) that has been observed among the rural population due to a number of factors. Despite the lower incidence of sedentary lifestyles compared to urban dwellers, there are other factors like higher levels of smoking in rural populations (Msyamboza KP, et. al., 2011), contaminated borehole water (Chimphamba, et. al., 2014), and high levels of pesticide exposure (Kosamu, et. al., 2020) that are resulting in increasing burdens of NCDs among the rural population.

Accessibility to timely diagnostics and medical management remains a continuing challenge in developing countries, more especially for populations in rural settings. These challenges are further compounded by high patient burdens and low human resources compliment. Consequently, clinical conditions are either missed at clinical presentation when in early pathological stages of development, or patients present late with well advanced disease states that are difficult and expensive to treat. As such screening and efficient clinical diagnostic services need to be made as accessible as possible.

In light of the above, rural populations, particularly pregnant women, would benefit from this innovative technology that would cut down on clinical review times, increase access to affordable and rapid screening and improve implementation of treatment or referral for further management.

 We would screen at a secondary health facility (Thyolo District Hospital in our proposed study) that receives patients from the community and primary levels for this study.  Once we have characterized disease profiles, we could determine how best to deploy as far into the communities as possible (Gov Malawi, 2022b).

References:

Chimphamba JB, Phiri OL, “Borehole water pollution and its implication on health on the rural communities of Malawi,” Malawi J Science and Technology 10(1), 2014

Government of Malawi (2022a) https://malawiplus.com/thyolo/

Government of Malawi (2022b) https://www.health.gov.mw/index.php/2016-01-06-19-58-23/national-aids

Kosamu I, Kaonga C, Utembe WA, “Critical review of the status of pesticide exposure management in Malawi,” Int J Environ Res Public Health. Sep 15;17(18):6727, 2020 doi: 10.3390/ijerph17186727. PMID: 32942751; PMCID: PMC7557847)

Msyamboza KP, Ngwira B, Dzowela T, Mvula C, Kathyola D, Harries AD, Bowie C, “The burden of selected chronic non-communicable diseases and their risk factors in Malawi: nationwide STEPS survey,” PLoS One. 2011;6(5):e20316. doi: 10.1371/journal.pone.0020316. Epub 2011 May 23. 2011 PMID: 21629735; PMCID: PMC3100352

Peterson I, Kapito-Tembo A, Bauleni A, Nyirenda O, Pensulo P, Still W, Valim C, Cohee L, Taylor T, Mathanga DP, Laufer MK. “Overdiagnosis of malaria Illness in an endemic setting: A facility-based surveillance study in Malawi,” Am J Trop Med Hyg. May 3;104(6):2123-2130. 2021 doi: 10.4269/ajtmh.20-1209. PMID: 33939628; PMCID: PMC8176516).

The Solution Design Team (Drs. John Robertson and Ryan Senger)

 Drs. John Robertson VMD PhD (Medical Pathology) and Ryan Senger PhD (Chemical Engineering) invented the Rametrix® technology (computational methods, devices, specimen spectral databases) described in this proposal.  Beginning with an idea (“Raman spectroscopy could be used to analyze biological fluids”) in 2014, they set out to prove this. Building on limited information on Raman spectroscopy of normal urine present in the medical and analytical literature, they profiled over 2500 urine specimens from healthy volunteers, patients with a variety of genitourinary and systemic diseases, dialysis patients, and even dogs at risk for developing bladder cancer.  They found:

• The composition of urine from healthy volunteers (no evidence of renal disease) was remarkably similar one specimen to the next, even when there were differences in sex, diet, and age of the volunteers,
• The composition of urine from the same individual did not vary over days, weeks, and months, although the diet of the person, physical activity of the person, and (females) hormonal cycles varied,
• The urine of patients with UTDs had ‘Raman spectral molecular fingerprints’ quite distinct from the ‘Raman molecular fingerprints’ of healthy volunteers – and that these ‘fingerprints’ (representative of hundreds of discrete molecules in urine) allowed screening of diseases.

They invented (with their research colleagues) novel ways to statistically analyze data (now using AI and computational genomic methods).  They designed and built devices to automate specimen analysis, making the technology available and scalable for mass screening of samples. 

Their efforts have provided a revolutionary method that uses simple, affordable, durable technology  that could provide disease screening to the global population facing the scourge of UTDs. 

The Solution Delivery Team (Drs. Mwayiwawa Madanitsa, Gama Bandawe, Andre Meulenaer, Penelope Muelenaer)

 Dr Madanitsa is a medical doctor and clinical epidemiologist who has worked in the public health system and has extensive experience in working with rural populations on clinical studies, particularly amongst pregnant women.  Through his experience, Dr Madanitsa has an acute appreciation of the burden and consequences of missed opportunities for the timely and effective implementation of clinical diagnosis and health care for communicable and non-communicable diseases, with an appreciation for the need and potential impact of innovative, low-cost technologies for the improvement of health outcomes and health care delivery.

Dr. Gama Bandawe is a medical virology PhD with a MSc in molecular and cell biology and a BSc in microbiology and biochemistry. He is currently a senior lecturer and head of the Department of Biological Sciences in the Academy of Medical Sciences at the Malawi university of Science and Technology. The Biological Sciences Department currently runs two recently developed bachelor’s programs which are the first of their kind in Malawi. These are BSc programs in Medical Microbiology and in Immunology. The BSc in immunology program has graduated 2 cohorts to date and these have already had a significant impact on the laboratory and diagnostics landscape in Malawi as the recent graduates took center stage in manning the national response to the COVID-19 pandemic. The department is currently in the process of rolling out two masters programs, one in Medical Microbiology and one in Infection and Immunity which will both be enrolling their first cohorts in November of this year. Dr Bandawe has 14 years of working experience in public health and biomedical research, laboratory management and academia. He has technical expertise in a variety of laboratory based and bio-statistical analytical techniques as well as bioinformatics related to virology, microbiology, molecular biology and immunology. Dr Bandawe has experience working with public health laboratory systems in South Africa and Malawi and has been involved in project management, coordination and administration in both settings. Dr Bandawe is a National Diagnostics Subcommittee member under the Health Cluster subcommittee of the Presidential Taskforce on COVID-19 (March 2020 – present) and a member of the Global Implementation Subcommittee of the Massachusetts General Brigham Health COVID Innovation Centre (June 2020 – present). He is currently Malawi University of Science and Technology Research Ethics Committee, Chairperson (March 2019 – present). Dr Bandawe’s current research interests include AMR surveillance and virus discovery with a focus on etiology of undiagnosed febrile illness. 

Andre Muelenaer, MD, MS, is a pediatric pulmonologist, and Penelope Muelenaer, MD, MPH, specializes in pediatric infectious diseases.  Both have been involved in global health for 40 years, beginning in 1982 when they were assigned to the Frankfurt Army Regional Medical Center (FARMC) in Germany.  As such they received pediatric patients sent from military and diplomatic missions for consultation, therapy and transport to US-based medical facilities.  During, and after their fellowship training, they remained in the US Army Reserve, gaining experience in global health through military training and service.  In 1990 they were activated for the First Gulf War.  Penelope Muelenaer served at Fort Bragg, North Carolina, where she educated soldiers about communicable diseases they might encounter in the Middle East.  Andre Muelenaer was deployed with the 401st Military Police, Enemy Prisoner of War, unit that was located in remote Saudi Arabia.  He was responsible for the acute healthcare, preventive medicine, food inspection, and emergency transport for 3500 US Army soldiers.  Additionally, he worked with leadership at a field hospital unit, co-located with the 401st, that provided acute and chronic healthcare services for the 22,000 Iraqi prisoners and refugees who arrived within a 48-hour period in late February 1991.  One of his primary responsibilities was to ensure that this vulnerable population of men was cared for in compliance of the Articles of the Geneva Convention.  Under his leadership, there were no deaths or disease outbreaks during the 4 months that these 22,000 men were detained.  For this, he was awarded the Army Commendation and the Bronze Star medals for service. 

In February 2004, Penelope was offered the opportunity to engage in a program to deliver and distribute food aid during the famine in Malawi.  As part of this 2-week program implementation trip, she was also asked by CitiHope International, an international humanitarian relief and development organization, to survey the needs for diagnosis and treatment of infants and children with HIV/AIDS.  She returned in October 2004, and worked in pediatrics at Mzuzu Central Hospital for almost two months as she gained additional insights regarding care for HIV infected children.  She lived in the local community, and was immersed in the culture as part of this experience.  Over the last eighteen years she has returned to Malawi, primarily to educate healthcare personnel there, and students from Virginia Tech. Of note is that in 2013 she completed her capstone project for her MPH in public health education in Malawi.  Following the principles of community based participatory research, she recruited thirty-six (36) households from an area surrounding Mzuzu, randomized them to three different forms of household water purification, and conducted water testing and surveys to determine the best means and acceptance rates of purification.  She returned in 2014 to share results of her study, and to provide additional education in the community in collaboration with local healthcare educators.  She has continued to work within the continuum of care, from household to tertiary care hospitals in Malawi to bring appropriate, affordable innovations created at Virginia Tech to the underserved.

Following a similar path as Penelope, Andre was deployed to Kyrgyzstan in February 2004 as part of Operation Provide Hope, a US Department of State program.  He conducted a series of educational sessions, related to treatment of asthma, that complemented delivery by CitiHope International of thousands of bronchodilator and steroid metered dose inhalers.  He was responsible for providing up to date information to 115 rural family physicians who had not received medical education since the fall of the Soviet Union in 1991.  Following this 2-week trip, he became the chair of the medical advisory committee for CitiHope International, and over several years of service became its chief medical officer.  In 2009, as the Chief Medical Officer he was tasked to conduct needs assessments for donation of medications, medical supplies, and medical equipment for a program supported by the Coca Cola Africa Foundation, ShareHope.  ShareHope was a collaboration between CitiHope and Medshare, an Atlanta based organization that provides surplus medical supplies and equipment to communities in need around the world. He conducted these assessments in hospitals throughout Africa, and learned very early that he could not assess the true needs unless he communicated with the true stakeholders; the physicians, nurses, patients, and families who lived and worked in these communities. 

As Drs. Muelenaer worked with individuals and communities in Malawi, they discovered that there were many individuals like them from communities in southwestern Virginia. These included university faculty and students, church groups, civic organizations, and the general public.  In September 2015 they organized a meeting of eleven people, and TEAM Malawi was formed. TEAM Malawi [Technology-Education-Advocacy-Medicine] is a transdisciplinary collaboration, based upon a community wellness model of health, designed to meet the challenges of resource-limited environments through community based participatory research/design/pedagogy.  Its Vision:  Utilize a community wellness model as the framework to integrate the activities of a disparate group of young investigators and mentors.  Its Primary Goal:  Student Engagement, with a desire to:  Foster global perspectives of students. Stimulate ideas for improving conditions in a developing country.  Create sustainable relationships that promote transdisciplinary models for development. Encourage evidence-based applied scholarly research.  Over the last seven years we have established working relationships with several universities in Malawi, conducted research projects in Malawi, mentored multiple senior design projects based upon needs in Malawi, and enriched the education of dozens of engineering students through activities at Virginia Tech and in Malawi.  The UNICEF sponsored African Drone and Data Academy, located at Malawi University of Science and Technology finds its origins in TEAM Malawi, as does the new library building being constructed at Mzuzu University after a fire completely destroyed their library in December 2015.

Our most recent activity was to visit Malawi in June 2022 with the mission being to foster relationships among key stakeholders in the government, healthcare facilities, and Malawi University of Science and Technology, as we bring technologies with the potential to impact healthcare.  At the Ministries of Health and Education, we established clear lines of communication, and developed a deep understanding of governmental policies and processes that will facilitate introduction of medical devices and healthcare IT beneficial to Malawi’s people.  Discussions at Kamuzu Central, Queen Elizabeth Central, and Thyolo District Hospitals provided us with information regarding high priority diseases, recommended locations for services, and basic needs for medical equipment and supplies.  All three hospitals welcomed services and research activities that could enhance their delivery of quality healthcare.  As evidenced by this proposal, Malawi University of Science and Technology has embraced the TEAM Malawi approach.  

Drs. Muelenaer have become a part of these communities, teach and practice the principles of community-based participatory research and design, and are dedicated to empowering the faculty and students at MUST and its surrounding communities to create wellness. 

This is a technology solution that is “shovel-ready”  for deployment and clinical research/investigation in a model low- and middle-resource environment - Malawi.  A strong, experienced Team is in place to lead this effort. In a nutshell, all that is needed funding to produce the technology and to initiate/run the demonstration project in Malawi.

The most significant barrier to overcome is a financial barrier. The entire cost of this demonstration project is being requested from The Challenge.  DialySensors Inc. will bear all personnel costs for DialySensors personnel without seeking funding from the proposed grant.  All other costs will be met by funds from the Challenge grant and there will be absolutely no costs/financial commitments/burdens in Malawi. 

• DialySensors Inc. will manufacture three, small, single-sample, portable analyzer systems (each - $40,000 – at cost of manufacturing and no profit) and one large, 50-sample hospital clinical lab analyzer ($60,000 – at cost of manufacturing and no profit), a total of $180,000;
• Delivery of the devices to points-of-care in Malawi is estimated between $15,000 - $25,000 (packaging, shipping, air transit, overland local delivery)
• A supply of consumables (glass sample vials, urine specimen collection equipment) for supporting collection and analysis of 25,000 patient and healthy human urines would be supplied and with the systems.  The estimated cost of these supplies is $40,000.
• Salary reimbursement for a full-time Study coordinator, skilled nursing/technical healthcare personnel (part-time), and faculty consultants (part-time) in Malawi to collect and analyze samples and conduct the study (four to six persons – one dedicated each system deployed at separate points-of-care) is estimated to be $100,000-150,000
• Patients and healthy human volunteers would be recruited and incentivized to participate.  In the first two years of this project, we would target the collection and analysis of 25,000 individual samples.  We estimate the costs associated with recruitment/enrollment and sample collection (incentivized) would be  $75,000

We do not anticipate any technical barriers.  We (DialySensors Inc.) have conducted the types of clinical research and investigation studies proposed here for the past five years in the US, using the equipment and technologies proposed here.  We have robust, agile manufacturing capabilities to produce and support this durable technology. 

There are going to be logistical barriers.  We plan to deploy our innovative solution (essentially, a point-of-care system for urinary tract health and disease screening in rural Malawi.  We know, from our interactions in Malawi over the past 10 years, that we can anticipate four logistical challenges.  First, the supply of electrical power (needed to run the analyzers) is undependable and intermittently interrupted.  To overcome this logistical challenge, we will supply each analyzer with solar and battery power – so that the analyzers are ready when patients need screening.  Second, access to the Internet is undependable and intermittently interrupted.  To overcome this logistical challenge, each analyzer and its computer will be designed for stand-alone operation, not needing connection to the Internet for access to programs, applications, or databases/data library (the needed databases will be present on each system). Intermittently, updating of a central database can occur when Internet veracity is assured, or data from patient files can be uploaded on thumb drives and physically transported for download in the Malawi National Raman Spectrum database.  Third, we know that equipment breaks.  To meet this logistical challenge, we will train student engineers and technicians at MUST (Malawi University of Science and Technology) to repair the equipment.  As the design of the systems is modular, swap out-replacement is simple.  A ‘spare set’ of all modular components (including a replacement Raman spectrometer and several probes), housing and drives, will be kept at MUST – ready for equipment servicing, as needed.  Fourth, each analysis requires a glass vial.  While in the US, these items are considered consumables, this is not the case in Malawi.  Working with engineering students are MUST, we will create a process to clean and reuse the glass vials, eliminating the need for costly and time-consuming replacement. We also will supply a backup stock of 25,000 vials to cover breakage and to prevent delays at the POC in urinalysis screening. 

There may be modest/minimal legal and regulatory barriers.  We will seek legal approval from the Food and Drug Administration to export our devices and technologies to Malawi with permitting granted for “Clinical Investigation/Research Use ONLY”. By the same token, we will seek approval from the Ministry of Health and Government of Malawi to import and deploy this technology for “Clinical Investigation/Research Use ONLY”. 

We do not anticipate significant cultural barriers.  Members of our Team in Malawi will be entirely responsible for the appropriate use of the technology in their healthcare environment.

Successful deployment and ‘proof-of-concept’ use of the Raman molecular chemometric urinalysis (Rametrix®) technology will create market opportunities for DialySensors Inc. worldwide.  

In low- and middle-resource countries, the technology could be provided “at-cost” to sponsoring governments and/or non-governmental organizations (NGOs).  Continued operation, data collection, and data analysis would need to be supported by governments and NGOs to provide sustained local autonomy and decision-making from data. 

In other venues (North America, Europe, South and East Asia, Australia, for example), there would be significant market opportunities for deploying this simple technology (Raman molecular chemometric urinalysis (Rametrix®)) in ‘point-of-care’ (POC) healthcare decision-making.  

An example may help illustrate the potential opportunity:

• Just as in Malawi and Sub-Saharan Africa, urinary tract infections (UTIs) are common in long-term care (LTC) facilities in the US,
• Just as in Malawi and Sub-Saharan Africa, prompt detection and appropriate therapy for LTC patients is key to management,
• Just as in Malawi and Sub-Saharan Africa, early detection of UTIs in LTC patients is problematic and cumbersome, requiring sample collection and transport, sample preparation, access to urine culture and urine cytology, laboratory equipment, and expertise – this is time-consuming and can be costly,
• In one US LTC patient study, the incidence of UTIs was as high as 7.43 per 1000 resident care days, according to the medical literature (Stevenson, et. al., 2005),
• In LTC facilities, with the prevalence of asymptomatic bacterial colonization of the urinary tract (bacteriuria) higher than overtly symptomatic UTI. In non-catheterized residents, asymptomatic bacteriuria has been estimated at 18% to 57% for women and 19% to 38% for men (Genao, et. al., 2012), 
• According to the most recent comprehensive data available (2016-2018), nearly two million US citizens live in 44, 500 assisted care facilities (residential assisted living, nursing home, hospice) (CDC, 2021),
• In 2019, CMS (Medicaid/Medicare) Long-Term Services and Support (LTSS) expenditures totaled $162.1 billion in FY 2019, with institutional (assisted living and nursing home) services accounting for $67.1 billion (41.4 percent) of the total expenditure (CMS, 2019).

There would be substantial benefits (effective detection and management of UTIs, improved patient comfort and quality-of-life, prevention of disease progression/complications) to placing and supporting Raman molecular chemometric urinalysis (Rametrix®)) systems in every LTC facility (and hospital) in the US (nearly 50,000 sites, combined).  The cost of purchasing and operating these systems would be borne by Medicaid/Medicare (CMS), private healthcare insurers, and foundation/clinic operators – generating a modest profit for DialySensors Inc. that could subsidize/reduce costs for low- and middle-resource environment systems and for ongoing research and technology evolution.

References: 

CDC, 2021 https://www.cdc.gov/nchs/fastats/nursing-home-care.htm; https://www.cdc.gov/nchs/npals/Survey-methodology-document03152021.pdf 

CMS, 2019 https://www.medicaid.gov/medicaid/long-term-services-supports/downloads/ltssexpenditures2019.pdf). 

Genao L, Buhr GT, “Urinary tract infections in older adults residing in long-term care facilities,” Ann Longterm Care. Apr;20(4):33-38. 2012 PMID: 23418402; PMCID: PMC3573848  

Stevenson KB, Moore J, Colwell H, Sleeper B, “Standardized infection surveillance in long-term care: interfacility comparisons from a regional cohort of facilities,” Infect Control Hosp Epidemiol. 26(3):231–238) 2005 

Current methods to evaluate urinary tract function and health are, and have been, insufficient to detect and manage urinary tract disease in low- and middle-resource environments, such as Malawi (and even long-term care facilities in the US).  The lack of appropriate, simple, resource/environment-appropriate technology, and lack of access to technology and healthcare, has burdened these countries and environments with potentially manageable diseases that may progress (predictably) to serious, chronic, and unmanageable disease, leading to significant morbidity and mortality.

We believe our solution and approach (Raman molecular chemometric urinalysis (Rametrix®)) can change this.  Why do we think it is innovative and catalytic?

• There are few, if any, simple, cost-effective devices or technologies available – either commercially available or as research tools - for profiling many different urinary tract diseases and urinary tract metrics (glomerular filtration rate – GFR, for example);

• We can profile the function and health of the urinary tract with a single 5 ml sample of urine, collected at the ‘point-of-care (POC)’;
• There is no physical or chemical processing needed to analyze the sample – it can simply be poured into a glass tube and scanned (less than one minute time and no requirement for highly-trained personnel to perform the operation);
• Data analysis (comparison of the patient sample with a robust local sample reference database/data library) provides meaningful, actionable information to healthcare providers within 15 minutes;
• The database/data library used for analysis is generated locally – at the site of use (Malawi, for example).  It is not reliant on published literature values that may be derived from distant patient populations with diverse (and potentially irrelevant) characteristics (race, environment, age, diet, among many others);
• Every patient or volunteer sample analysis improves the database/data library by adding locally-important information.  As the size and complexity of the local database/data library increases (days-months-years), important health trends can be followed and analyzed temporally.  One example of this is our (US) COVID19 (SARS-Cov-2) database/data library.  We collected and analyzed urine samples from healthy volunteers (no evidence of urinary tract disease) for ‘baselining’ between 2016-2018.  When COVID19 appeared in the US (2020) we had a completely ‘non-exposed’ group of reference spectra (healthy volunteers, 2016-2018) to compare with urine Raman spectra from persons with COVID19 disease.  We soon discerned a unique molecular fingerprint of COVID19 disease (not the virus or viral antigens – but molecules indicating metabolic and inflammatory reactions to infection (Robertson, et. al., 2022).  The local database/data library is a powerful and important tool for long-term detection and monitoring disease trends;
• The simplicity of data collection (small amounts of voided urine) facilitates repetitive sampling of the same patient – during treatment and over time.  Rametrix® analysis of patient samples over time allows evaluation of treatment efficacy, can rapidly signal a lack of response to treatment (this is very important in the use of antibiotics for management of UTIs), and provides actionable information to healthcare providers over days-weeks-months-years.  We recently reported (PDF, attached paper in review, Peer J) that Rametrix® analysis of urine from patients with diabetes mellitus could differentiate patient with diabetic kidney disease from patients with diabetes who had other forms of kidney disease (and who would be treated with different medications and management) (Kavuru, et. al., submitted, 2022);
• The simple analytical device(s) are easily manufactured, populated with low-cost, commercially-available Raman spectrometers, are self-contained for local data capture and analysis, are modular in construction (components easily and quickly swapped out and replaced), are engineered for predictable operation at ambient temperature in challenging environments (heat, cold, humidity), and with minimal power needs (standard 120V, battery, solar panel sources).  They literally have been designed to work dependably at any POC, at any time;
• The devices and technology can be easily operated and maintained by minimally-trained personnel (from high school/technical school students to nurses, laboratory technicians, and physicians). 

References:

Kavuru, V, Senger, RS, Robertson, JL, Choudhury, D, “Analysis of urine Raman spectra differences from patients with diabetes mellitus and renal pathologies,” submitted,Peer J 

Robertson JL, Senger RS, Talty J, Du P, Sayed-Issa A, Avellar ML, et al. “Alterations in the molecular composition of COVID-19 patient urine, detected using Raman spectroscopic/computational analysis,” PLoS ONE 17(7): e0270914. https://doi-org.ezproxyberklee.flo.org/10.1371/journal.pone.0270914 2022

Impact Goal #1: Provide a low-cost, simple analytical system for assessing urinary tract health and for urinary tract disease screening in a low-resource environment.  Why?  Urinary tract diseases in the target population (residents in rural Malawi) are common and are under-diagnosed/treated due to a lack of point-of-care (POC) medical analytical resources.  With the use of our novel Raman molecular chemometric urinalysis (Rametrix®) system, caregivers at the POC will quickly (15-30 minutes) have actionable medical analytical information for decision-making (Examples: “Does the Raman data indicate that patient may have diabetes and diabetic kidney disease?”  Or “Does the Raman data indicate the patient may have an acute urinary tract infection and what is the likely cause (organism)?”  Or “Does the Raman data indicate the patient may have bladder cancer?”).

Impact Goal #2: Facilitate medical decision-making at the POC.  How? Providing on-site caregivers with actionable medical analytical information will allow them to decide how best to treat their patients.  This may involve using more definitive diagnostic testing (imaging, laboratory studies – which can be costly and difficult to provide at the POC), instituting empirical treatment (e.g., prescribing antimicrobial therapy), or referral to specialty facilities and providers (again, this may be costly and difficult to provide, requiring transit over long distances). Also, the ability to provide actionable medical analytical information at the POC will allow caregivers to assess the efficacy of treatments and periods of observation and to longitudinally profile patient health – at a cost of pennies to gain, store, and retrieve the information.  This will empower both patients and entire healthcare systems – where deployed and used. 

Impact Goal #3: Tailor medical information to the demographics of the target patient population and conditions of the patients’ lifestyle.  How and Why?  Each and every patient and volunteer sample can and will be used to create databases and data libraries unique to the low- and middle-resource countries and regions in which the samples are collected.  These critical reference tools will quickly incorporate hundreds ->many thousands of analytical data points that can be ‘mined’ with artificial intelligence (AI) tools to define health and disease at a patient, location, or regional/national level.

Impact Goal #4: Improve the health and quality-of-life (QOL) in low- and middle-resource countries and regions.  Why?  There are many reasons, but among them:

• Access to effective healthcare alleviates suffering of patients and their families,
• Access to affordable healthcare improves the QOL on a national and regional level,
• No person should be denied healthcare because of a lack of resources or the system to provide those resources,
• Early screening and effective intervention in many urinary tract health problems will slow or prevent the predictable progression to more serious disease problems that are costly or difficult to manage,
• Not knowing prevalent diseases in low- and middle-resource countries and regions prevents development and essential metrics/strategies for controlling these diseases,
• From an epidemiological perspective – the health of the people of The World is interconnected.  Recent history is replete with examples – SARS-CoV1, SARS-CoV2, influenza, monkeypox– and the consequences for not screening and promptly acting on actionable medical analytical information.  A convincing argument was made that the Global COVID19 (SAR-CoV2) Pandemic directly and indirectly affected all of the Seventeen (17) Sustainable Development Goals of the United Nations (UN, 2022),
• From an economic perspective, there is a direct linear relationship between access to healthcare, improvement in individual and workforce productivity and subsequent reduction in poverty, decreased pediatric and maternal morbidity and mortality, foreshortened lifespan, and premature death (UN, 2022).

Reference:

UN, 2022 https://www.un.org/sustainabledevelopment/health/

According to the United Nations (Sustainable Goal #3) “Ensuring healthy lives and promoting well-being at all ages is essential to sustainable development.”  Our four Impact Goals directly address this UN challenge and focus.

Key indicators for measuring progress toward our goal of creating, deploying, and using a novel and innovative technology (Raman molecular chemometric urinalysis (Rametrix®) system) in the context of low- and middle-resource environments (Malawi) will include: 

• First, acceptance, critical review, and meaningful follow-up on this proposal submission by MIT/Gate SOLVE; response to critical review (DialySensors Inc.), followed by funding of the proposal, fabrication and deployment of three systems in Malawi, training of key medical and support personnel;
• Second, development, acceptance, and execution of clinical study protocols at three study sites;
• Third, collection and analysis of patient samples; reporting of actionable medical analytical information to caregivers; followed by patient outcomes assessments;
• Fourth, evaluation to see how patient outcomes are affected by the presence of this novel technology;
• Fifth, developing interactions and integration of the technology and data with Malawi One Health Surveillance Program(s); and with the activities of the appropriate district health management team(s).

We already have achieved the first four of these indicators/metrics in the US, in the past four years.  This is shown by publications on the technology (Rametrix®) and use of the technology in evaluation of patient health/disease.  These publications are referenced earlier in this proposal.  We have shown our technology can accurately assess renal function (GFR), detect and stage chronic kidney disease, detect complications of diabetes mellitus, detect complications of COVID19 disease, and screen for the presence or absence of bladder cancer.  With the support of DialySensors Inc. and MIT/Gates, we will achieve similar indicators and metrics in Malawi, showing that this novel technology can be readily deployed and effectively used in low- and middle-resource environments.

Reference: https://www.un.org/sustainabledevelopment/health/

The diffusion of innovations theory of change will be applied for this project.  A two stage, formal and informal, approach will be undertaken by the research team.  First, the new innovation will be formally presented by Dr Mwayiwawo Madanitsa, a trusted opinion leader with shared beliefs and norms that define the social structure in the medical community of the Thyolo District Hospital.  Healthcare providers who ordinarily evaluate and treat patients will be asked to attend these formal educational sessions. These sessions will include the basic science behind Raman molecular chemometric urinalysis, current clinical applications that have evolved from research in the USA, and the rationale for focusing on acute and chronic urinary tract disease for this study.  The procedure for obtaining specimens for use by the research team will be reviewed.  Next, the study coordinator, who has training and experience in medical laboratory procedures and public health education, will visit each healthcare provider, enabling informal exchanges of information.  It is anticipated that early adopters will influence change among their peers, reinforced by continued presence of the study coordinator.

Within the context of the Five Stages of the Innovation-Decision Process, accelerated adoption is anticipated through practical knowledge as to the underlying principles of Raman molecular chemometric urinalysis and how it works.  Users will be persuaded by heightened awareness of the advantages of this approach to screening for disease, and greater insight in terms of expected outcomes.  Decisions to accept the innovation should follow more quickly, resulting in implementation of Raman molecular chemometric urinalysis.  Confirmation, or commitment to use will be constantly reinforced through frequent communication by the research team members.   

Reference: 

Rogers, EM. Diffusion of Innovations (5th ed.) New York, NY: Free Press. 2003

The core technology (Raman spectroscopy) has been extensively described previously.  A summary follows:

“Raman spectroscopy is a method for determining the molecular composition of liquids (such as urine) and solid materials (powder mixtures, for example).  Irradiation of the specimen (urine, for example) with laser energy, deforms/stretches/bends/vibrates the chemical bonds of thousands of different molecular components in the specimen.  As soon as the laser irradiation is stopped, these chemical bonds return to their resting state, while releasing small amounts of bond deformation stored energy.  This energy release (photons) can be detected at different wavenumbers and can be used to create a Raman spectrum that represents the ‘molecular fingerprint’ of the liquid.  Comparing one Raman spectrum from a healthy person, for example, with the Raman spectrum from a patient with suspected disease, allows detection of differences in ‘molecular fingerprints’ that characterize diseases (see below). 

Our technology consists of three components: an analytical device containing sample holders and a Raman spectrometer, computational algorithms to process raw Raman spectra to useful information, and a series of reference databases that contain a library of Raman spectra from historical samples.  The reference database spectra are the product of analysis of thousands of urine specimens from healthy volunteers and from patients with diseases that affect the urinary tract specifically and the body, as a whole.  It is very important to understand that the reference databases/data libraries are built “on site” when the samples are analyzed – allowing comparison of samples/spectra from local populations.  These databases and data libraries grow and grow, as more specimens are added, eventually expected to contain the results of analysis of hundreds of thousands of specimens, representing the regional population and temporal conditions.  Data interpretation is not reliant on historical datasets, collected from different populations (North America and Europe, for example), that have been historically biased against datasets from low- and middle-resource countries.

The process of analysis is very simple!  Patients urinate into a cup and some of the urine (1-5 ml) is then transferred into a reusable glass tube that is inserted into a holder in the analyzer.  The analyzer lid is closed and the laser is then activated (up to 15 times) and the raw Raman spectrum produced electronically, by detecting of the Raman scatter radiation.  This electronic spectrum is then computationally normalized (to eliminate artifacts) and these result are sent electronically for comparison with other reference Raman spectra in the database/data library.  Artificial intelligence algorithms are used to find particularly prominent and important differences between a single patient specimen and the reference databases/data library, as well as searching for trends and differences between many types of samples (male vs female, young vs old, healthy vs diseased, acute disease vs chronic disease, and so forth). 

A single patient urine sample (5ml) can be screened for the presence of infection, acute kidney injury, connective tissue disease, diabetic kidney disease, chronic kidney disease, bladder inflammation, and urinary tract cancer (see References, following this text section). The analysis costs pennies per measurement (consumables) and results are available within15-30 minutes (done, as previously stated, by electronic comparison of patient spectral results with reference database of >2000 spectra). The technology can quantify glomerular filtration rate (GFR), proteinuria, and hematuria in a single sample.  It has been extensively validated. References cited above.”

As previously noted, healthcare providers at the point-of-contact (POC) will collect clinical samples from consented patients and analyze them with our technology. They will be incentivized with government, non-governmental, and supplemental (DialySensors Inc.) funding. Aside from a financial incentive, local healthcare providers have a vested interest in, and duty to attend to the health and well-being of patients in their local communities.

Members of this proposal team (Robertson, Senger, A Muelenaer, P Muelenaer) all hold faculty positions at Virginia Tech (VT).  Drs. Robertson and Senger are the co-founders of DialySensors Inc., an approved and supported ‘faculty start-up’. Their company (DialySensors Inc.) has an exclusive license from VT and Virginia Tech Intellectual Properties for commercializing (their) faculty-initiated technology.  As members of the VT Community, all participates ascribe to and have pledged to support Virginia Tech’s Principles of Community (https://www.inclusive.vt.edu/about/vtpoc.html).  This states:

“Virginia Tech is a public land-grant university, committed to teaching and learning, research, and outreach to the Commonwealth of Virginia, the nation, and the world community. Learning from the experiences that shape Virginia Tech as an institution, we acknowledge those aspects of our legacy that reflected bias and exclusion. Therefore, we adopt and practice the following principles as fundamental to our on-going efforts to increase access and inclusion and to create a community that nurtures learning and growth for all of its members:

• We affirm the inherent dignity and value of every person and strive to maintain a climate for work and learning based on mutual respect and understanding.
• We affirm the right of each person to express thoughts and opinions freely.
• We encourage open expression within a climate of civility, sensitivity, and mutual respect.
• We affirm the value of human diversity because it enriches our lives and the University.
• We acknowledge and respect our differences while affirming our common humanity.
• We reject all forms of prejudice and discrimination, including those based on age, color, disability, gender, gender identity, gender expression, national origin, political affiliation, race, religion, sexual orientation, and veteran status.
• We take individual and collective responsibility for helping to eliminate bias and discrimination and for increasing our own understanding of these issues through education, training, and interaction with others. 

We pledge our collective commitment to these principles in the spirit of the Virginia Tech motto of Ut Prosim (That I May Serve). 

As an organization, DialySensors Inc. has two employees (both VT-affiliated) who also ascribe to the Principles of Community.  Mr. Amr Sayed Issa is a biomedical engineer and also a recent refugee from Syria.  Ms. Lacey Ngo is a part-time research associate and full-time undergraduate student in the College of Engineering.  Other members of the DialySensors Inc. Team include COO Richard Daugherty (a retired engineer)(retired VT faculty), IT Officer Pang Du (current VT faculty), Medical Advisor Allan Sklar MD (current VT faculty), Business Consultant WT Lee (current Carilion Clinic employee), and Regulatory Consultant   Mehmet Kosoglu (former VT Engineering student and FDA employee).  We are proud of the diversity of our DialySensors Inc. Team.

DialySensors Inc. is an incorporated (C-Corp) for-profit company in Virginia.  The company develops technology (devices, computational software), based on their proprietary Raman spectroscopy platform (Rametrix®), to screen for, and provide actionable medical analytic information for human healthcare.  The business model is based on technology sales to healthcare providers and their insurers, including health maintenance organizations.  As well, the company licenses their technology and leases their equipment. 

The entire ‘product line’ of technology is focused on disease management and monitoring with molecular urinalysis.  With this focus, the patient population targeted and served are patients with kidney disease/failure and urinary tract neoplasms.  Given the disproportionately high prevalence of diabetes mellitus and hypertension in economically-disadvantaged communities in Virginia and the US, DialySensors Inc. has fostered relationships in these communities.  As an example, DialySensors Inc. recently collaborated with faculty at the Virginia Commonwealth University Center for Pharmacy Practice and Innovation and CrossOver Ministry, to begin the introduction of our technology (precisely analogous to the current proposal) for detection and management of urinary tract disease in diabetic patients in an economically-disadvantaged area in and around Richmond, VA. The goal of our work in these communities is to detect urinary tract health problems, provide actionable medical analytical information to healthcare providers, and to assist them in determining efficacious management that controls disease progression. 

Services (devices, sample analysis) are provided through support of clinical investigation grants from governmental and non-governmental sources.  Several technology products (drug analyzers, volatile substance analyzers) will be marketed in 4Q22 to non-profit and for-profit healthcare organizations, as well as the US Dept. of Defense.  These analyzers and technologies will be used to detect dilution/diversion of controlled substances, medication formulation errors, and the composition of exotic fuel mixtures (obviously all separate applications – using the same Raman technology platform).

DialySensors Inc. will continue to fund its R&D and clinical work with a combination of donations and grants (State of Virginia, National Institutes of Health, Dept. of Veterans Affairs, and Dept. of Defense).

Additionally, DialySensors Inc. will sell custom-built analyzers for detection of controlled substance diversion (ProofMed™ Scanner V1.0), analysis of volatile/non-volatile mixtures (AVProof™ Scanner V1.0), and patient molecular urinalysis (Rametrix® AutoScanner V2.0).  With these sales, there will be ongoing revenues in terms of maintenance contracts and database/software licensing. 

We have and will continue to seek investment capital to support R&D and clinical work, as well as ongoing device and IT structure development.

We have been successful in obtaining funds to support DialySensors R&D, clinical investigations, and product development.  Some examples:

• “Dialysensing™: Improving the efficacy and patient outcomes of hemodialysis and peritoneal dialysis through the use of Raman spectroscopy and multivariate statistical analysis,” Commonwealth Commercialization Fund $58,000 (2015)
• “DialySensors: Re-defining the management of renal failure in dialysis patients,” Robertson, J, Senger, R, VT KnowledgeWorks Innovation Fund $100,000 (2016-2017)
• “Improving decision-making in organ transplantation,” 4-VA Collaborative Research Grant $25,000 (2020-2021)

• ” Rametrix® AutoScanner V2.0 automated molecular urinalysis system,” VTIP POC Fund $40,000 (2021)
• “Rametrix® ProofMed V1.0 ™: Point Of Care (POC) management and reduction of potential Controlled Substance (CS) diversion in hospital settings,” Virginia Innovation Partnership Corporation $75,000 (2022)
• “COVID19 detection using Raman molecular urinalysis,” Virginia Bioinformatics Institute private donor $20,000 (2022)
• “Detecting renal complications associated with COVID-19 infections using Raman molecular urinalysis (RMU),” Carilion Clinic Research Acceleration Fund $75,000 (2022)
• Private Investor Funds $250,000 (2019, 2022)