Heterogeneous vancomycin intermediate Staphylococcus aureus (hVISA) strains are well-known worldwide clinical problem. These strains are associated with vancomycin treatment failure. In India, increasing incidences of hVISA has been observed. Macro E-test method is easy to screen for early detection of hVISA and Cu2O-NPs can be used as effective antibiofilm agents in diverse medical and household devices

hVISA Trends in India
Professor Kashi Nath Prasad and his student Dr. Avinash Singh observed higher vancomycin minimum inhibitory concentration (MIC) among methicillin-resistant Staphylococcus aureus (MRSA) at Sanjay Gandhi Post Graduate Institute of Medical Sciences (SGPGIMS), Lucknow (India). They were worried about the higher vancomycin MIC at SGPGIMS, because cases of vancomycin treatment failures with higher vancomycin MIC, have been observed by them against MRSA. Vancomycin is normally used to treat MRSA type infections but its overuse causes the development of vancomycin-intermediate S. aureus (VISA) and heterogeneous VISA (hVISA). They thought hVISA has emerged at their hospital. The identification of hVISA was very labour intensive and a tricky task. They took the help of Professor Kechi Hiramatsu from Jutandu University in Tokyo, Japan for a positive control and standardizing the protocol for hVISA detection. Dr Avinash started screening for hVISA strains for three consecutive years and the data was really shocking. He noticed that hVISA is not only emerging at northern India but incidences are increasing steadily from 4.5% to 6.7%. He checked the patient records and found that few cases were non-responsive to vancomycin treatment and patients finally succumbed to these strains. It was really alarming because vancomycin was supposed to be the best and cost-effective drug for the treatment of MRSA infections.
They published their data in the Journal of Microbial Drug Resistance. In an institutional Continued Medical Education (CME) they have convinced the clinicians to consult the Department of Microbiology of SGPGIMS if they notice vancomycin is not giving a normal response. CME created more awareness among intensive care unit (ICU) doctors and lead them to find ways to identify hVISA. The gold standard test for hVISA detection is Population Analysis Area under curve (PAP-AUC) but it is time consuming (takes 3-4 days) and labour intensive. ICU doctors need quick results to start appropriate antibiotics in the case of hVISA related sepsis and related infections. So, an alternative rapid method was urgently needed for hVISA detection.

Creating a Method for Rapid Screening of hVISA
Dr. Prasad and Dr. Avinash reviewed previous research and studied the physical and molecular properties of these strains in the quest for rapid diagnostic marker for hVISA. They observed a molecular marker, accessory gene regulator I (agr I) which was common in 80% of hVISA isolates at their center. But reports from previous studies showed different agr patterns in strains from other countries. Most of the hVISA isolates were slow grower and have long lag phase in their growth cycle and had lesser generation time in hVISA isolates. Dr Avinash found highest positive predictive value for quick identification of hVISA in 24-48 hours by “Macro E-test method”. Dr Avinash and Dr Prasad were little disappointed and like other researchers who were doing work in other countries, they were not able to get any rapid marker for hVISA detection but they concluded that if hVISA is screened by macro E test method and have type 1 agr gene clinicians should start giving higher doses of Vancomycin or other alternatives antibiotics depending upon the patient condition. Dr Avinash published above mentioned study in Journal of Global Antimicrobial Resistance and Indian Journal of Medical Microbiology. Their group was the first in India who performed such large-scale screening of hVISA. Their findings were also acknowledged by Indian Council of Medical Research (ICMR) in New Delhi, India.

Alternative treatment for hVISA type Infections
During those days, Dr. Gajbhiye (Chemistry Professor) from Indian Institute of Technology at Kanpur, was synthesizing novel Cuprous Oxide (Cu2O) nanoparticles and wanted to see its antimicrobial effects against bacteria. He sent a proposal to Dr. Prasad and his group for exploring antimicrobial properties of Cu2O nanoparticles. Dr. Avinash took the challenge and started screening the antimicrobial properties of novel copper nanoparticles and results were really promising. They found that these Cu2O nanoparticles were showing significant antibacterial properties against Gram positive bugs compared to Gram negative bugs. At the same time, they were trying to explore the resistance mechanism of hVISA and they found cell wall thickness of hVISA isolates played a major role in their resistance. Due to cell wall thickness, vancomycin is blocked and cannot reaches its target. They planned to test Cu2O nanoparticles against these bugs. Surprisingly, they found Cu2O nanoparticles were very effective in killing hVISA isolates without any toxic effects at used concentration. They also explored the mode of action of Cu2O nanoparticles by performing a couple of experiments. They found that these nanoparticles worked by disrupting the cell membrane and causes the leaking of intracellular material of the bacteria and ultimately leading to bacterial death. They published their findings in the American Society of Microbiology’s Antimicrobial Agent and Chemotherapy. Presently they are working on developing cuprous oxide coated household and medical devices for hospital use.

Lessons Learned
Ultimately, from this experience, I learned that you should dream big, take challenges, make good and smart plans, do the hard work, don’t shy away from trying new things and don’t worry about failures. With a passion for research and perseverance, researchers will definitely achieve their goals.

Dr. Avinash Singh wants to pursue his research dream in India and choose Indian Institute of Technology, Mandi for further research. He is working on developing biomarkers for early diagnosis of sepsis and exploring the dissemination mechanism of antibiotic resistance in pristine locations of India.

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