The Way It Was.

Mary Osborn was a native Californian. She was an undergraduate at the University of California, Berkeley, where she worked in the laboratory of I.L. Chaikoff. She received her PhD at the University of Washington, where her work on the role of folic acid coenzymes in one-carbon metabolism revealed the mechanism of action of methotrexate. After postdoctoral training with Bernard Horecker in the Department of Microbiology at New York University (NYU), she embarked on her research career as a faculty member in the NYU Department of Microbiology and in the Department of Molecular Biology at Albert Einstein College of Medicine. In 1968 she moved as one of the founding faculty of the new medical school of the University of Connecticut, where she remained until her retirement in 2014. Her research was focused on the biosynthesis of the endotoxin lipopolysaccharide (LPS) of gram-negative bacteria and on the assembly of the bacterial cell envelope. She made seminal contributions in these areas. She was the recipient of numerous honors and served as president of several important scientific organizations. Later in her career she served as chair of the National Research Council Committee on Space Biology and Medicine, advisory to the National Aeronautics and Space Administration (NASA), which produced an influential report that plotted the path for NASA's space biology research program in the first decade of the twenty-first century. Dr. Osborn died on Jan. 17, 2019.

Transforming clinical microbiology with bacterial genome sequencing.

Whole-genome sequencing of bacteria has recently emerged as a cost-effective and convenient approach for addressing many microbiological questions. Here, we review the current status of clinical microbiology and how it has already begun to be transformed by using next-generation sequencing. We focus on three essential tasks: identifying the species of an isolate, testing its properties, such as resistance to antibiotics and virulence, and monitoring the emergence and spread of bacterial pathogens. We predict that the application of next-generation sequencing will soon be sufficiently fast, accurate and cheap to be used in routine clinical microbiology practice, where it could replace many complex current techniques with a single, more efficient workflow.

[Bibliometric analysis of bacterial quantitative proteomics in English literatures].

OBJECTIVE: To analyze the worldwide advances on bacterial quantitative proteomics over the past fifteen years with bibliometric approach. METHODS: Literature retrieval was conducted throughout the databases of Pubmed, Embase and Science citation index (SCI), using "bacterium" and "quantitative proteomics" as the key words. The deadline is July 2013. We sorted and analyzed these articles with Endnote X6 from the aspects of published year, the first author, name of journal, published institution, cited frequency and publication type. RESULTS: 932 English articles were included in our research after deleting the duplicates. The first article on bacterial quantitative proteomics was reported in 1999. The maximal publications were 163 related articles in 2012. Up till July 2013, authors from more than 23 countries and regions have published articles in this field. China ranks the fourth. The main publication type is original articles. The most frequently cited article is entitled with "Absolute quantification of proteins by LCMSE: a virtue of parallel MS acquisition" by Silva JC, Gorenstein MV, Li GZ, et al in Mol Cell Proteomics 2006. The most productive author is Smith RD from Biological Sciences Division, Pac. Northwest National Laboratory. The top journal publishing bacterial quantitative proteomics is Proteomics. CONCLUSION: More and more researchers pay attention to quantitative proteomics which will be widely used in bacteriology.

Updates in immunoassays: bacteriology.

There are many immunoassays available that provide rapid, accurate and sensitive results. The intent of this article was to provide a brief overview of some of the products and methodologies available for clinical use and to discuss some of the principles behind the methodology and instrumentation. In the area of infectious disease, the use of immunoassays ensures rapid turnaround times that will result in the administration of prompt, accurate treatment for the patient. Ultimately, this will improve overall patient outcomes while possibly decreasing the costs associated with increased hospital stay. In conclusion, immunoassays are essentially easy to perform, cost-effective, produce highly sensitive and specific results, and allow the medical laboratory professional the ability to report accurate results in a timely manner.

Global tuberculosis research and its future prospects.

Tuberculosis is the infectious disease that causes the most deaths each year in the world. Around 25% of the population is estimated to be infected with, Mycobacterium tuberculosis, the bacteria that gives rise to the disease, and more than one and a half million people die each year from this cause. A rigorous bibliometric analysis has been developed around tuberculosis disease, and the most remarkable results are presented in this paper. It is observed that interest in tuberculosis is growing, and the control of its spread has become one of the main health priorities in the world, with the United States, the United Kingdom, and India, leading the research in this area. On the other hand, it has been observed that there are two main health concerns around the tuberculosis: drug-resistant tuberculosis and co-infection with HIV. Finally, conclusions are offered, playing a frontline role in science policy decisions and research performance evaluations.

[The future of automation in bacteriology]. / Automatisation en bactériologie : quel avenir ? Un exemple concret dans le cadre d'une consolidation de laboratoires universitaires.

Bacteriology remained essentially manual for many years. After a partial automation for blood cultures, identifications and sensitivity testing, new technological developments including robotisation and digital pictures made it possible to open new ways. In the context of economic pression and need to increase the quality, automation offers multiple advantages concerning increase of productivity, standardization, traceability and decreasing of the delay to obtain the results. Moreover the use of digitalized pictures opens the way to tele-bacteriology, particularly useful when considering the merging of hospital laboratories because it makes it possible to geographically dissociate strict manipulation from the validation of the results and from the consultant activity of the microbiologist. The choice criteria of the equipment are detailed as well as the experience of the LHUB-ULB bacteriological laboratory which was automated at the time of merging of the Brussels public hospital laboratories and developed a conclusive experience of tele-bacteriology for the peripheral lab.

[Recent progress in mycobacteriology].

Mycobacterium tuberculosis is one of the most successful bacterial parasites of humans, infecting over one-third of the population of the world as latent infection without clinical manifestations. Over 8.8 million new cases and nearly 2 million deaths by tuberculosis (TB) occur annually. TB poses a significant health threat to the world population. The goal of this symposium is to open new avenues for combating tuberculosis. The speakers have presented their data and provided control strategies against tuberculosis and pulmonary disease due to M. avium complex (MAC) from aspects of molecular epidemiology, pathogenesis, serodiagnosis, new anti-TB drugs, and vaccine development. Drs. Maeda and Murase have reported that the 12-locus VNTR analysis is very useful for molecular epidemiology of M. tuberculosis strains isolated in Japan better than IS6110-RFLP and suggested that the analysis is powerful tool for the molecular epidemiology. Drs. Matsumoto and Kobayashi have discovered a protein, mycobacterial DNA-binding protein 1 (MDPl), overproduced in dormant M. tuberculosis that plays key roles in latent/ persistent infection, disease progression, and host protection. They have concluded that MDP1 may be a possible target for anti-tuberculosis drugs and vaccines. Drs. Kitada and Maekura have developed serodiagnosis of MAC disease based on enzyme immunoassay (EIA) by detecting anti-glycopeptidolipid (GPL) antibody in sera of human patients. GPL is specific for MAC. The EIA is a simple, rapid and accurate measure with high sensitivity and specificity. The levels of antibody also reflect disease activity. A large-scale clinical multicenter study is currently in progress. Dr. Makoto Matsumoto has discovered an innovative new anti-TB drug, OPC-67683 that is a derivative of nitroimidazole compounds. OPC-67683 inhibited mycolic acid synthesis and exerted potent antimycobacterial activity, including multidrug-resistant M. tuberculosis. Multidrug therapy using OPC-67683 could also shorten the course of chemotherapy. The drug is clearly the most promising new anti-TB agent that has been identified in many years. Dr. Okada has presented the vaccine candidates for TB, such as HVJ-liposome/HSP65 DNA+IL-12 DNA and HVJ-envelope/HSP65 DNA+IL-12 DNA. The candidates exhibited an excellent protective efficacy in mice compared to current BCG vaccine, and improved histopathologic lesions induced by M. tuberculosis infection. The candidates also exerted the therapeutic effect in mice against both drugsusceptible TB and extensively drug-resistant TB. Using the cynomolgus monkey model (similar to human TB), HVJ-liposome/ HSP65 DNA+IL-12 DNA provided higher protective efficacy than BCG assessed by mortality. The combination of BCG and HVJ-liposome/HSP65 DNA+IL-12 DNA by the prime-booster procedure could lead to a synergistic effect of 100% survival in infected monkeys. These data suggest that the novel DNA vaccine is a possible candidate for human clinical trials. This symposium has highlighted new advances in our understanding of molecular epidemiology and pathogenesis of "Mycobacteriology" and development of new serodiagnostics, anti-TB drugs, and vaccines. 1. The establishment of the quick genotyping method for TB in Japan using the variable numbers of tandem repeats (VNTR): Shinji MAEDA, Yoshiro MURASE (Research Institute of Tuberculosis, Japan Anti-Tuberculosis Association) The 12-locus VNTR analysis that we have established optimally for Mycobacteriun tuberculosis in Japan was superior to the proposed 15-locus VNTR method in European countries. The discriminatory power of our system was also higher than that of IS6110-based restriction fragment length polymorphism analysis. In future, we will investigate the stability of copy number in each locus by using the strains that suspected epidemiological links in contact investigations. 2. A virulence factor of Mycobacterium tuberculosis, which contributes to persistent infection, reactivation, and host protection: Sohkichi MATSUMOTO (Department of Host Defense, Osaka City University Graduate School of Medicine), Kazuo KOBAYASHI (Department of Immunology, National Institute of Infectious Diseases) Majority of adult tuberculosis is caused by reactivation of previously implanted Mycobacterium tuberculosis. During latent infection, some bacilli are in dormant state, which confers some survival advantage to not only bacteria but also the host. We presented that a protein overproduced in dormant M. tuberculosis plays key roles in persistent infection, disease progression, and host protection. We also presented utility of this protein, such as development of anti-tuberculosis drug and vaccine. 3. Serodiagnosis of Mycobacterium avium complex pulmonary disease by enzyme immunoassay using glycopeptidolipid antigen: Seigo KITADA, Ryoji MAEKURA (Department of Internal Medicine, National Hospital Organization National Toneyama Hospital) The diagnosis of Mycobacterium avium complex pulmonary disease (MAC-PD) and/or its discrimination from pulmonary tuberculosis is sometimes complicated and time consuming. We have developed serological test by enzyme immunoassay that detect serum antibody to glycopeptidolipid antigen. The serodiagnosis is useful for the rapid diagnosis of MAC-PD and differential diagnosis from pulmonary TB. The antibody levels reflected the disease activity including radiographic severity. 4. A novel antituberculous agent, OPC-67683: Research and development: Makoto MATSUMOTO (Microbiological Research Institute, Otsuka Pharmaceutical Co., Ltd.) We initiated a program to screen new antituberculous agents that have potential to shorten the total duration of treatment, provide improved efficacy against MDR-TB, be useful in treating HIV co-infected patients, and target latent TB infections. Our efforts led to the discovery of OPC-67683, a novel oxazo-imidazole derivative with a distinctive characteristic as a subclass mycolic acid inhibitor. Our evaluation studies confirmed OPC-67683 to possess potent in vitro and in vivo antituberculous activity, suggesting potential usefulness in alleviating the current TB problems. 5. The development of novel vaccines against M. tuberculosis: Masaji OKADA (Clinical Research Center, National Hospital Organization Kinki-Chuo Chest Medical Center) We have developed a novel tuberculosis (TB) vaccine (HVJ-liposome/ or HVJ-envelope/HSP65 DNA+ IL-12 DNA). The vaccine provided remarkable protective efficacy in mouse compared to BCG vaccine, and improved the histopathological tuberculosis lesions. This vaccine also exerted therapeutic effect in vivo against XDR-TB as well as drug-sensitive TB in mice. Furthermore, by using the cynomolgus monkey (similar to human tuberculosis), this novel vaccine provided higher protective efficacy (mortality) than BCG mortality. Furthermore, the combination of HSP65+IL-12/HVJ and BCG by the priming-booster method showed a synergistic effect in the TB-infected cynomolgus monkey (100% survival). These data indicate that our novel DNA vaccine might be useful against TB for human clinical trials.

[Recent advances in the field of oral bacteriology].

The oral cavity is inhabited by more than 600 bacterial species; these species compete for nutrients or coexist in order to survive along with the indigenous population. Extreme conditions are prevalent in the oral cavity, and these conditions are influenced by our immunity and variations in nutrition, temperature, and pH. Pathogens that cause dental caries or periodontal disease can survive in these extreme environments; these pathogens are virulent and can cause several diseases. Therefore, research on oral bacteriology is warranted to analyze the virulence factors of these bacteria as well as to ascertain environmental stress responses, interactions between bacteria and human immunity, comparisons of bacterial genomes, and oral microflora. In this review, we provide new data in the fields of bacteriology, immunology, and genomics and describe recent advances in the field of oral bacteriology.

[Current status of bacteriological studies at prefectural and municipal public health institutes in Japan].

Prefectural and municipal public health institutes are located in prefectures and ordinance-designated cities in Japan, and play a vital role in the regional surveillance of infectious diseases and foodborne illnesses. These institutes, in close cooperation with national institutes such as the National Institute of Infectious Diseases and the National Institute of Health Sciences, construct the national surveillance network for infectious diseases and their causative agents. Bacteriological examinations and studies on a variety of infectious diseases and foodborne illnesses are core activities of prefectural and municipal public health institutes, through which novel and important bacteriological findings have been acquired. In this article, we report the latest findings regarding bacteriological examinations/studies and interesting cases at these institutes, especially concerning foodborne illnesses, tuberculosis, and antimicrobial resistances.

Bacterial genomes for the masses: relevance to the clinical laboratory.

Bacterial genome sequencing has revolutionized the research landscape and promises to deliver important changes to the clinical microbiology laboratory, through the identification of novel diagnostic targets and through the birth of a new discipline or "genomic epidemiology." Current progress and future prospects for exploitation of genome sequences in clinical bacteriology are reviewed.

The mycobacteriology laboratory. Past, present, and future.

The role of the mycobacteriology laboratory in tuberculosis control programs has been underestimated in the past, but now laboratory services are proven to play an essential role if properly implemented. The importance and reliability of these services in the future will depend on their centralization in specialized mycobacteriology laboratories that will be able to put new technologies into practice. The shortest turnaround time and the greatest cost effectiveness can be achieved by direct submission of raw specimens to such laboratories, especially for the management of new tuberculosis patients.

[The bacteriology laboratory of tomorrow]. / Le laboratoire de bactériologie de demain.

The development of needs and the conditions for performing examinations in clinical bacteriology leads us to consider that bacteriological analysis should eventually: be closer to clinical requirements. This results in an accelerated bacteriology (whose response-times approach those of other biological specialties) and in a consideration of the diagnostic and therapeutic importance of the response; be less costly. This leads to an increase in productivity, and involves an introduction of computers, automatic systems for antibiograms and identification, and appropriate and inexpensive bioreagents; consider the requirements of the technicians in order to avoid diminishing the interest of the work. This development demands a recognition of needs, whether expressed or not, and a rejection of olds habits that are not adapted to the requirements of a bacteriology which is closer to the patient, quicker, and finally, less costly.

[Progress in clinical microbiology of Mycobacterium avium complex and prospect on its future].

Recent advance of technology in isolation and identification methods for mycobacteria, including BACTEC 460 TB System, polymerase chain reaction (PCR), DNA probe test, etc., enabled us to perform rapid and accurate identification of mycobacterial pathogens from various clinical specimens. Our recent study revealed usefulness of BACTEC 460 TB System, on the basis of the high efficiency in detecting the organisms in sputum specimens with much shortened recovery time, as compared to the Ogawa method. Moreover, accurate identification was possible, when the diagnostic system was combined with AccuProbe test. PCR using appropriate primers specific to Mycobacterium, M. tuberculosis complex (MTC), or M. avium complex (MAC) is useful to detect mycobacterial organisms directly in clinical specimens in a rapid manner. By this technique, in particular nested PCR, it is now possible to detect very small number of the organisms, as few as one bacterium, in test specimen, although it is still needed to resolve some difficulties due to contaminating inhibitors in the specimens. In our nested PCR system using Mav17 and Mav18 primers specific for Mycobacterium, especially MAC, we found that detectable degree of DNA amplification was possible even by replacing purified DNA sample, extracted from the organisms by considerably time consuming procedures, with bacterial suspension as a DNA template, by adding about 10(4) of organisms. The three DNA probe testings, Gen-Probe, AccuProbe and SNAP kits, are useful for rapid identification of MAC, MTC, M. kansasii and M. gordonae, and consistent results were obtained by these diagnostic kits. SNAP is able to distinguish the X component of MAC from the other two types of MAC.(ABSTRACT TRUNCATED AT 250 WORDS)

Broad-range PCR: past, present, or future of bacteriology?

PCR targeting the gene encoding 16S ribosomal RNA (commonly named broad-range PCR or 16S PCR) has been used for 20 years as a polyvalent tool to study prokaryotes. Broad-range PCR was first used as a taxonomic tool, then in clinical microbiology. We will describe the use of broad-range PCR in clinical microbiology. The first application was identification of bacterial strains obtained by culture but whose phenotypic or proteomic identification remained difficult or impossible. This changed bacterial taxonomy and allowed discovering many new species. The second application of broad-range PCR in clinical microbiology is the detection of bacterial DNA from clinical samples; we will review the clinical settings in which the technique proved useful (such as endocarditis) and those in which it did not (such as characterization of bacteria in ascites, in cirrhotic patients). This technique allowed identifying the etiological agents for several diseases, such as Whipple disease. This review is a synthesis of data concerning the applications, assets, and drawbacks of broad-range PCR in clinical microbiology.

[New antituberculosis vaccines]. / Nouveaux vaccins antituberculeux.

Today, only one type of vaccine is available to protect against tuberculosis. This vaccine, called Bacille Calmette-Guérin (BCG) was developed approximately 100 years ago and has been administered at least 3 billion times. Initial multicenter studies have indicated an up to 93% efficacy against childhood tuberculosis mortality. Subsequently, many studies on BCG efficacy have been carried out, with highly variable results, ranging from 0 to 90% efficacy. The reasons for this heterogeneity are not well understood. Large clinical studies have shown that booster vaccinations with BCG do not improve the BCG efficacy. Therefore, new vaccines are urgently needed. Today, there are essentially two lines of efforts being pursued in several laboratories. One of them aims at replacing BCG with superior vaccines. This strategy focuses either on improving existing BCG by constructing strains that overproduce certain protective antigens and/or by improving its immunogenicity, or on starting anew by genetically attenuating virulent Mycobacterium tuberculosis. The second line of research aims at adding onto BCG vaccination, such as a heterologous prime-boost strategy. For this strategy BCG is used as a first vaccine, followed by individual antigens as a booster. These antigens can be presented in several different ways, and preference is currently given to the so-called latency antigens. Both approaches have yielded encouraging results in animal models, and some of them have now entered clinical trials. Although still far from human applications, it is hoped that these strategies will ultimately help to reduce the enormous burden that is caused by tuberculosis.