Samir P. Singh

BACKGROUND: The epidemiology of tuberculosis in urban populations is changing. Combining conventional epidemiologic techniques with DNA fingerprinting of Mycobacterium tuberculosis can improve the understanding of how tuberculosis is transmitted. METHODS: We used restriction-fragment-length polymorphism (RFLP) analysis to study M. tuberculosis isolates from all patients reported to the tuberculosis registry in San Francisco during 1991 and 1992. These results were interpreted along with clinical, demographic, and epidemiologic data. Patients infected with the same strains were identified according to their RFLP patterns, and patients with identical patterns were grouped in clusters. Risk factors for being in a cluster were analyzed. RESULTS: Of 473 patients studied, 191 appeared to have active tuberculosis as a result of recent infection. Tracing of patients' contacts with the use of conventional methods identified links among only 10 percent of these patients. DNA fingerprinting, however, identified 44 clusters, 20 of which consisted of only 2 persons and the largest of which consisted of 30 persons. In patients under 60 years of age, Hispanic ethnicity (odds ratio, 3.3; P = 0.02), black race (odds ratio, 2.3; P = 0.02), birth in the United States (odds ratio, 5.8; P < 0.001), and a diagnosis of the acquired immunodeficiency syndrome (odds ratio, 1.8; P = 0.04) were independently associated with being in a cluster. Further study of patients in clusters confirmed that poorly compliant patients with infectious tuberculosis have a substantial adverse effect on the control of this disease. CONCLUSIONS: Despite an efficient tuberculosis-control program, nearly a third of new cases of tuberculosis in San Francisco are the result of recent infection. Few of these instances of transmission are identified by conventional contact tracing.

BACKGROUND: In the United States there have been recent outbreaks of multidrug-resistant tuberculosis. These outbreaks have primarily involved persons infected with the human immunodeficiency virus (HIV). METHODS: We collected clinical information on 17 patients seen at a New York City hospital who had repeatedly positive cultures for Mycobacterium tuberculosis. Analysis of restriction-fragment--length polymorphisms (RFLPs) was performed on serial isolates of M. tuberculosis obtained from these patients. RESULTS: Six patients had isolates that remained drug-susceptible, and the RFLP patterns of these isolates did not change over time. Eleven patients had isolates that became resistant to antimicrobial agents. The RFLP patterns of the isolates from six of these patients remained essentially unchanged (two strains showed one additional band) despite the development of drug resistance. In five other patients, however, the RFLP patterns of the isolates changed dramatically at the time that drug resistance was detected. The change in the RFLP pattern of the isolate from one patient appeared to be the result of contamination during processing in the laboratory. In the remaining four patients, all of whom had advanced HIV disease, the clinical and microbiologic evidence was consistent with the presence of active tuberculosis caused by a new strain of M. tuberculosis. CONCLUSIONS: Resistance to antituberculous drugs can develop not only in the strain that caused the initial disease, but also as a result of reinfection with a new strain of M. tuberculosis that is drug-resistant. Exogenous reinfection with multidrug-resistant M. tuberculosis can occur either during therapy for the original infection or after therapy has been completed.

Various techniques have been adopted to impart a biological responsiveness to synthetic hydrogels for the delivery of therapeutic agents as well as the study and manipulation of biological processes and tissue development. Such techniques and materials include polyelectrolyte gels that swell and deswell with changes in pH, thermosensitive gels that contract at physiological temperatures, and peptide cross-linked hydrogels that degrade upon peptidolysis by cell-secreted enzymes. Herein we report a unique approach to photochemically deform and degrade disulfide cross-linked hydrogels, mitigating the challenges of light attenuation and low quantum yield, permitting the degradation of hydrogels up to 2 mm thick within 120 s at low light intensities (10 mW/cm(2) at 365 nm). Hydrogels were formed by the oxidation of thiol-functionalized 4-armed poly(ethylene glycol) macromolecules. These disulfide cross-linked hydrogels were then swollen in a lithium acylphosphinate photoinitiator solution. Upon exposure to light, photogenerated radicals initiate multiple fragmentation and disulfide exchange reactions, permitting and promoting photodeformation, photowelding, and photodegradation. This novel, but simple, approach to generate photoadaptable hydrogels portends the study of cellular response to mechanically and topographically dynamic substrates as well as novel encapsulations by the welding of solid substrates. The principles and techniques described herein hold implications for more than hydrogel materials but also for photoadaptable polymers more generally.

Molecular strain typing by restriction fragment length polymorphism analysis was used to demonstrate that two clusters of Mycobacterium tuberculosis cultures involving six patients resulted from cross-contamination in the mycobacteriology laboratory. Contaminated cultures were processed by the decontamination procedure and were read on the BACTEC instrument following acid-fast bacillus smear-positive specimens from patients with active tuberculosis. Investigation of these episodes suggested opportunities for modification of laboratory procedures to minimize cross-contamination and confirmed the adverse medical and public health consequences of false-positive cultures. Strain-typing results were used in decisions regarding patient care, including the curtailment of unnecessary treatment in one patient. Molecular strain typing appears to be a valuable means of identifying false-positive cultures of M. tuberculosis in selected settings.

To address the challenges associated with defined control over matrix properties in 3D cell culture systems, we employed a peptide functionalized poly(ethylene glycol) (PEG) hydrogel matrix in which mechanical modulus and adhesive properties were tuned. An HT-1080 human fibrosarcoma cell line was chosen as a model for probing matrix influences on tumor cell migration using the PEG hydrogel platform. HT-1080 speed varied with a complex dependence on both matrix modulus and Cys-Arg-Gly-Asp-Ser (CRGDS) adhesion ligand concentration, with regimes in which motility increased, decreased, or was minimally altered being observed. We further investigated cell motility by forming matrix interfaces that mimic aspects of tissue boundaries that might be encountered during invasion by taking advantage of the spatial control of the thiol-ene photochemistry to form patterned regions of low and high cross-linking densities. HT-1080s in 100 Pa regions of patterned PEG hydrogels tended to reverse direction or aggregate at the interface when they encountered a 360 Pa boundary. In contrast, HT-1080s were apparently unimpeded when migrating from the stiff to the soft regions of PEG peptide hydrogels, which may indicate that cells are capable of "reverse durotaxis" within at least some matrix regimes. Taken together, our results identified matrix regimes in which HT-1080 motility was both positively and negatively influenced by cell adhesion or matrix modulus.