Saturday, December 25, 2010
Wednesday, December 22, 2010
Immunochromatographic strip test for detection of genus Cronobacter

Tuesday, December 21, 2010
MIT Researchers Reconstruct Evolution of 3 Billion-Year-Old Microbes

Micro- and nanocantilever devices and systems for biomolecule detection

Monday, December 13, 2010
DNA Sequencing Matches Cholera Strain in Haiti with Bacteria from South Asia

A team of researchers from Harvard Medical School, Brigham and Women's Hospital, and Massachusetts General Hospital, with others from the United States and Haiti, has determined that the strain of cholera erupting in Haiti matches bacterial samples from South Asia and not those from Latin America. These findings, which appeared in the New England Journal of Medicine, conclude that the cholera bacterial strain introduced into Haiti probably came from an infected human, contaminated food or other item from outside of Latin America.
To identify the probable origin of the cholera strain in Haiti, scientists used a third-generation, single-molecule DNA sequencing method developed by Pacific Biosciences. They determined the genome sequences of two Haitian cholera samples and three cholera samples from elsewhere around the world. Based on advanced imaging technology, the method enables researchers to observe a natural enzyme synthesizing a strand of DNA in real time. As such, the technology actually tracks and documents nature at work, a rapid approach compared to other sequencing technologies. The method allowed a comprehensive analysis and comparison of critical DNA features among the various cholera samples, which included single nucleotide variations, insertions and deletions of particular portions of the genome, and structural variations. The analysis showed a close relationship between the Haitian samples and the seventh pandemic variant strains isolated in Bangladesh in 2002 and 2008.
Genetic changes occur quickly, within hours in the lab and probably weeks within the environment, through natural modes of DNA swapping and mutation among bacteria. Their evolution is based, in part, on the acquisition, loss, and alteration of mobile genetic elements, including DNA from the CTX bacterial virus, which bears the genes encoding the cholera toxin, and other genetic sequences that may make a particular strain more adapted to a given ecosystem. The resulting heterogeneity has been used to categorize strains of the seventh pandemic and to understand their transmission around the globe.
Wednesday, December 8, 2010
UTHealth professor to receive service award from American Society for Microbiology

Known nationally for her research into single-cell organisms that affect oral health, Millicent "Mimi" Goldschmidt, Ph.D., a professor of microbiology and molecular genetics at The University of Texas Health Science Center at Houston (UTHealth), has been selected to receive the 2011 American Society for Microbiology (ASM) Founders Distinguished Service Award. The award will be presented at the ASM General Meeting Awards Banquet and Dinner in New Orleans on May 22.
"Dr. Goldschmidt has furthered the understanding of the basic microbiology of the mouth," said Larry R. Kaiser, M.D., president of UTHealth. "She has done an exemplary job of serving her professional community, her scientific community and her teaching community." Microbiology is the study of cells that are invisible to the naked eye. These tiny cells include bacteria, viruses and fungi. Goldschmidt studies microbes involved with dental decay, gum disease and oral cancer, as well as rapid methods of detection (biosensors, microarrays and nanoparticles).
Before joining the UTHealth faculty in the early 1970s, Goldschmidt was the coordinator of the protocol to plan the biological tests that would be employed in the lunar receiving laboratory on the first returned moon rocks. She was also instrumental in developing isolation protocols for Apollo astronauts returning from the moon, which ensured that infectious organisms would be detected and contained.
When she started her professional career in Texas five decades ago, Goldschmidt said there were really no rapid methods to detect microorganisms. Her research contributed to the development of rapid immunological and biosensor types of detection methods to pinpoint salmonellae, E. coli and oral microbes. She consults and lectures nationally and internationally on biosensors, microarrays and nanoparticles.
Sunday, November 28, 2010
A mobile phone app that detects STDs!

Tuesday, November 23, 2010
Fung’s Double Tube Leads Way to Quicker Pathogen Detection

The Fung Double Tube method is relatively easy to implement. The system uses one large tube and one small tube. Insert the small tube into the larger tube that holds a water sample. Then add a specially melted agar – a gelatin-like product used for solidifying culture media as a thickening agent – to create a thin layer between the two tubes that will grow C. perfringens. The unit is then placed in an incubator at 42 degrees C.
“The system makes anaerobic microbiology very simple,” Fung said. “One can see a C. perfringens colony in the Double tube in about four to five hours. We can know in five to six hours how many living colonies of C. perfringens per milliliter of water there are. That is a major improvement and it’s so cheap.”
The system works well for water testing, but more research is necessary to prepare it for use in the food industry. Fung’s research team is examining how to apply it for use with ground beef so the meat won’t have to be incubated overnight before pathogen counts can be obtained.
More Accurate Diagnostic for Influenza and Respiratory Syncytial Virus Using Nanoparticle Probes

Monday, November 15, 2010
A New Read on DNA Sequencing

Wednesday, November 10, 2010
WIll FDA develop a Guidance on RMMs?

Friday, November 5, 2010
Automating the Micro QC Lab (Part 4)
Michael - Anyone who has attempted to validate a rapid solution knows the hurdles that must be overcome. Should companies be concerned that automation adds another level of complexity to the validation?
Steve – Not necessarily. In actuality, this could just be the opposite. A rapid method solution like ours or others in the market can greatly simplify the validation process, as automation could remove a number of validation performance requirements that other alternative methods might require. Additionally, automated methods that are based on compendial testing may also simplify this process. For most compendial microbiology methods, we utilize a certain sample size, media, incubation time and temperature and acceptance limits. Using a rapid method that has similar (or the same) testing elements will be straightforward to validate, since the difference is the automation. Methods that deviate and require different or additional steps may introduce variables and risk factors that will need to be addressed in the validation program.
Thursday, November 4, 2010
New Publication on Rapid Sterility Testing

Wednesday, October 27, 2010
USP Discussion on RMMs and recommendations for USP Chapter 1223
The final session of this year’s PDA Global Conference on Pharmaceutical Microbiology is focused on the direction that the USP will take with the existing informational chapter 1223, Validation of Alternative Microbiological Methods. Questions were provided to the meeting attendees and here is an excerpt of some of the comments and responses:
Question: Has USP 1223 Validation of Alternative Microbiological Methods been useful or an impediment for the selection, validation and implementation of RMMs?
Response 1: Yes, the chapter has been useful as providing guidance for RMM validation.
Response 2: Yes, but we would like to have more guidance on sensitivity and limit of detection strategies. For example, methods for developing very low inoculum levels, such as 1 cell.
Response 3: The use of statistics for each validation criteria should be more clearly defined and relevant.
Response 4: Much work has been done on limit of detection for sterility testing and these methods are appropriate for use and should be incorporated into the USP guidance.
Response 5: There needs to be a good balance between specific guidance, such as acceptance criteria, and background information into the reasons why the guidance is provided. The chapter should not be a white paper.
Response 6: We need a simple benchmark on how to validate RMMs. The food industry and AOAC have addressed this 20 years ago!
Question: Do you like to see examples?
Response 1: In the EU, regulators took the example in Ph. Eur. 5.1.6 as gospel and that caused many problems for companies validating RMMs. As a result, the next revision of 5.1.6 will not contain an example but will be published in PharmEuropa.
Response 2: I want to see real, practical and successful case studies of how RMMs have been validated, not theoretical examples.
Response 3: Recommendation is not to have an example but to put this information in Pharmacopeial Forum and/or reference other guidance documents that will provide examples, such as PDA Technical Report #33.
Response 4: I want to see guidance on controls based on technology platforms and applications (e.g., negative and positive controls). But don’t provide specific examples, but more suggestions on good controls that should be run.
Question: Should compendial guidance documents (JP, USP, and EP) on the validation of RMMs be harmonized?
Response: By a show of hands, many in the audience said yes.
Final summary statements from the USP based on these discussions:
We may be making this much harder than it should be. We need to go back and take a closer look at our recommendations in this chapter and provide a more “user-friendly” set of recommendations. Because there are many different processes and products that would utilize RMMs, it is really up to you, the users of RMMs, to define how to best validate these new systems. USP intends to provide better guidance on the use of alternative methods with input from stakeholders.
USP 1223 needs to be revised. There needs to be a clarification of sensitivity, limit of detection and limit of quantification, with specific validation criteria and not sole reliance on parallel testing. The use of CFUs is difficult because there is no good quantitative definition of what a CFU is. Statistical models used with validation criteria needs to be revisited. Microorganisms chosen during validation should be appropriate for the intended applications, process and product. We need to consider how to handle slow growing microorganisms, and address the use of appropriate controls.
Next, it appears that specific examples should not be included to avoid regulatory expectations that may not be appropriate. Instead, maybe we should reference PDA TR #33 and/or submit a Stimuli Article. Any reference to specific RMM technologies should not be included.
We should address the relevance of referee tests for short shelf-life products such as biologics.
We should work toward harmonizing all RMM guidance documents; however, this may be easier said than done. This is not a simple matter and could be time consuming. However, we will discuss how we can interact with other organizations to determine if our revisions can take into account other guidance documents.
Finally, we need to have more discussions with the stakeholders of RMMs more frequently than we have in the past, and we will discuss how we can seek out mechanisms to make this happen.
Japanese PMDA Perspectives on RMMs

FDA Perspectives on RMMs


Regulatory RMM Perspectives at the PDA Global Microbiology Conference: The Australian TGA

Vivian Christ, Australian TGA first reviewed some of the policies and guidance that they follow with regard to RMMs. The TGA utilizes relevant sections in the Ph. Eur. and BP in that these compendia allow for the validation alternate methods. They also rely on the validation guidance from USP 1223, Ph. Eur. 5.1.6, PDA TR #33, and ISO 17025 (validation of non-standard methods), to name a few. From the legislative perspective, the TGA turns to the TGA GMPs, which allows for other acceptable methods as long as they are shown to be equivalent to those in the GMP guide, as well as Annex 11 (computer validation) and Annex 15 (IQ, OQ, PQ). However, unlike other regulatory agencies, such as the FDA, the TGA only “quietly” embraces new technologies but they have not come out with a formal statement or policy.
The TGA views RMMs to be used in a wide range of applications, including finished product testing and in-process testing. Validation expectations include a DQ, IQ and OQ performed by the vendor, and the PQ (jointly performed by the vendor and user) and verification of the method using actual product. Testing on potential interfering substances should be performed, and for users intending to develop a matrix testing strategy (i.e., grouping products together), a justification for doing so should be provided. Furthermore, there is the expectation that equivalence or superiority to classical method is demonstrated, as well as the computer validation of software. For a qualitative method, the validation should address specificity, sensitivity, ruggedness, robustness, and equivalence. For quantitative tests, the user should demonstrate accuracy, precision, ruggedness, robustness, equivalence sensitivity, linearity, and specificity.
Many companies have discussed implementing RMMs with the TGA but very few have actually taken the plunge and have moved forward. The TGA doesn’t really understand the reason for this. However, when companies do come in to discuss their intentions, the TGA encourages them.
TGA does have some regulatory concerns. For example, what happens if a company obtains a positive result by a RMM but not the referee test (what does this mean?). Next, do specifications need to be changed, and how will the company use the results from RMM testing, such as batch release? All of these points must be considered. From an administrative perspective, the local inspectors are starting to see RMMs but primarily for in-process testing, where there is no requirement for a formal regulatory change submission. Additionally, there is no published policy on the testing requirements for RMMs. From an economic standpoint, the TGA is considering cutting the costs associated with formal change submissions and RMMs, such as lumping many products together for a single RMM. Hopefully, we will hear more about this in the near future. In summary, the TGA hopes that more companies will move forward, validate and implement RMMs.
Tuesday, October 26, 2010
Pharmacopoiea Perspectives on RMMs Provided by USP, Ph. Eur. and JP Expert Committees
Rapid Method Session 2 at the PDA Global Microbiology Conference
Rapid Method Session 1 at PDA Global Microbiology Conference

Jennifer Gray of Novartis Pharma AG, Switzerland, presented their strategy for validating the Millipore Milliflex Rapid as an alternative ATP bioluminescence RMM to the traditional compendial sterility test. The drivers for a rapid sterility test included the early identification of product contamination events, a reduction of through put time for sterile drug product release, and to increase the company’s level of expertise in the field of rapid microbiological methods. Novartis validated a 5-day sterility test using 22 heat-stressed cultures (7 ATCC strains and 15 environmental isolates). The cultures were also used to determine the most optimal medium to be used in the system. The FDA approved comparability protocols outlining the validation strategy for multiple products, EMA approval was obtained in February 2010 and MHRA approval was obtained in May 2010.
Amelia Tait-Kamradt, Pfizer, discussed their assessment of Pall’s new GeneDisc system. They conducted a number of studies addressing specificity, limit of detection, ruggedness, robustness, and the impact that excipients may have on the ability of the system to detect microorganisms. More information on the GeneDisc system may be found on our website on the Technology page.
Dr. Geert Verdonk of Merck presented his validation studies using the Charles River Laboratories Endosafe PTS. Dr. Verdonk explored the use of this rapid and portable endotoxin detection system as part of Merck’s PAT-RMM program. A variety of validation studies were also presented.
Monday, October 25, 2010
Dr. Ed Tidswell Discusses Viable but Non-Culturable (VBNC) Organisms
FDA's Hussong Discusses Objectionable Organisms and RMMs

Dr. David Hussong, FDA CDER, reviewed the impact that B. cepacia can have on pharmaceutical product and the reasons why this organism may be considered objectionable for certain dosage forms. He stated that current test methods, including finished product testing, may not be sufficient in detecting objectionable organisms, and that it is more critical to look for these types of organisms during in-process screening using rapid methods.

Opening Session at PDA Global Microbiology Conference
Friday, October 22, 2010
Live RMM Blog Next Week!

Remember to follow my live blog from the PDA Global Conference on Pharmaceutical Microbiology next week!
On Tuesday and Wednesday I will blog LIVE from the PDA Micro meeting during the rapid micro methods sessions. Topics will include technology updates, regulatory perspectives (from US, EU and Japanese regulators), and the current Pharmacopoeia positions by experts from the USP, Ph. Eur. and JP.

Monday, October 18, 2010
New Pharmaceutical Microbiology Textbok Includes RMMs
Injectable product manufacturing is booming because of the growth of new biopharmaceuticals and small molecule anticancer drugs. The requirements for contamination control will become even more stringent than today. Isolators or blow-fill-seal equipment have already replaced the conventional clean rooms and LAF-hoods in many production facilitities. Conventional microbiological monitoring methods, requiring 3 to 5 days of incubation will become inappropriate. Equipment is already available allowing real time, simultaneous viable and non-viable counting.
This book is a useful reference guide for the SMB (Small and Medium Business) pharmaceutical sector which does not have the resources to have access to such top-quality information in this field. This book therefore represents an unparalleled and unprecedented text in the field of pharmaceutical and medical device microbiology. Perhaps even more outstanding is the fact that this book not only covers subject matter and technical content which is established as best and expected practice, but also includes content regarded as possible, future and emerging technology or processes.
The results of 45 years of scientific and technological development are laid down in these 33 chapters. These chapters, all written by international experts, give a vivid picture of today’s pharmaceutical microbiology. The high standard of the chapters makes it an essential reference guide that should be on the shelf of everyone who is involved or interested in this field.
Wednesday, October 6, 2010
LIVE RMM BLOGGING During the PDA Global Microbiology Conference
Three day Microbiology GMP and RMM Conference in Istanbul, Turkey, Dec. 13-15
Sunday, October 3, 2010
Participants needed for online survey on RMM return on investment
High-throughput Universal Probe Salmonella Serotyping (UPSS) by nanoPCR
A new publication from the September 2010 Journal of Microbiological Methods discusses the use of nano liter PCR to detect Salmonella. This is quite interesting opportunity in automating and miniaturizing PCR procedures. Here is the abstract:
Salmonella enterica subsp. enterica serovar identification is of great importance with respect to outbreak monitoring and case verification. Therefore rapid, sensitive and cost efficient detection of Salmonella spp is indispensable within microbiology labs. To amalgamate single tube isolate identification with Salmonella typing, we developed the high-throughput Universal Probe Salmonella Serotyping (UPSS) technique based on nano liter PCR. In comparison to the classical approach, where O- and H-antisera are applied, the UPSS relies on specific gene content amplification of Salmonella spp. by an universal TaqMan assay for all markers and identification of the specific amplicon pattern. To enable high-throughput technology we employed a chip format containing 1024 wells loaded by an automated liquid-handling system which allowed us to perform TaqMan PCR reactions in volumes of 100 nano liters per well. Herein we present proof of principle of the UPSS method by the use of a test panel of 100 previously serotyped Salmonella isolates to successfully verify the usability, accuracy and feasibility of the newly developed UPSS approach. We found that the methodology of the UPSS technology is capable of unequivocally identifying 30 Salmonella serotypes on a single chip within 3 hours but can be highly parallelized by the use of multiple PCR machines. Therefore the UPSS method offers a robust and straightforward molecular alternative for Salmonella detection and typing that saves expensive chemistry and can be easily automated.