Nancy E. Dunlap

A ‘safe-site’ for Salmonella typhimurium is within splenic polymorphonuclear cells☆

Following oral or systemic infection with Salmonella typhimurium, the focus of infection is in the liver and spleen. The majority of Salmonella surviving in the liver and spleen by 4 h post infection are already in an environment where they are largely protected from subsequent killing. Previous studies have shown that the majority of surviving Salmonella are intracellular. In the present study we sought to determine the cell type containing most of the cell-associated Salmonella liberated from the spleen. We enriched for Salmonella-containing cells by Ficoll-Hypaque separation followed by fluorescence-activated cell sorting. Approximately 85% of the total intracellular Salmonella were found in Mac-1+/J-11d+ cell fractions of the Ficoll-Hypaque band and pellet. By microscopic examination of stained cells from the sorted cell populations, it was evident that virtually all of the Salmonella were in polymorphonuclear cells (PMN). The numbers of Salmonella observed microscopically were similar in numbers to Salmonella colony forming units detected by plating. Salmonella containing PMN in the Ficoll band generally contained a single bacterium, while those from the probably less healthy cells in the Ficoll pellet generally contained several Salmonella.

Immunology of tuberculosis

TB is a chronic, necrotizing infection caused by M. tuberculosis. The clinical manifestations of disease are the result of a balance between the host response and bacterial virulence. Cellular immunity is responsible for effective control of infection, but cytokines released during the process of cellular immunity may also cause harm to the host. Humoral immunity plays little part in protection against TB. Individuals with defective cellular immunity are much more susceptible to disease from M. tuberculosis and are more likely to have a disseminated form of TB.

Molecular Differentiation of Mycobacterium tuberculosis Strains without IS6110 Insertions

By using standard restriction fragment length polymorphism, 6 zero-copy IS6110 Mycobacterium tuberculosis isolates were identified from 1,180 Maryland isolates as part of the National Tuberculosis Genotyping Surveillance Network Project. By using various genotyping methods, we demonstrated that this zero band cluster can be differentiated into six genotypes.

IS6110 insertions in Mycobacterium tuberculosis: predominantly into coding regions.

We read with interest the recent publication by Benjamin et al. ([2][1]) regarding the characterization of IS 6110 insertion sites in the direct repeat (DR) region of Mycobacterium tuberculosis . This topical and relevant study described the dissection of a single molecular event leading to an

Long-Term Molecular Analysis of Tuberculosis Strains in Alabama, a State Characterized by a Largely Indigenous, Low-Risk Population

ABSTRACT With a tuberculosis case detection rate of 5.9 per 100,000 population in 2001, Alabama ranked twelfth highest in the United States. However, cases among foreign-born and human immunodeficiency virus-infected individuals remain low in Alabama. To understand the endemic statewide disease pattern, tuberculosis strains were studied for clustering in a long-term population-based study from January 1994 to May 2000. IS6110 restriction fragment length polymorphism analysis was performed for 1,834 strains. Spoligotyping was used as a secondary typing method for the 37% of isolates displaying a restriction fragment length polymorphism pattern with <6 IS6110 copies. A total of 721 (41%) patients provided isolates that composed 114 clusters, each containing isolates from 2 to 136 patients, suggesting that recent transmission accounted for 35% of tuberculosis cases. Demographic, behavioral, and clinical characteristics of patients with clustered versus nonclustered isolates stratified by low-copy-number strains (<6 IS6110 copies) versus high-copy-number strains (≥6 IS6110 copies) were evaluated. Younger age, black race, a history of alcohol abuse, and homelessness were predictors of clustering of low-copy-number, strains and younger age, urban residency, alcohol abuse, homelessness, noninjection drug use, and a history of incarceration and/or cavitary disease were predictors of clustering of high-copy-number strains. By identifying local characteristics of tuberculosis clustering through molecular fingerprinting, control programs can distribute their limited resources to impact the transmission of tuberculosis in high-risk populations and evaluate strain distribution across geographical areas.

Childhood tuberculosis in Alabama: epidemiology of disease and indicators of program effectiveness, 1983 to 1993.

An 11-year review of childhood tuberculosis in Alabama was made in order to define indicators of program effectiveness in interrupting community transmission. Minority (nonwhite) children, 96% of whom were black, had the highest risk of disease (odds ratio, 5.5; 95% confidence interval, 3.9, 7.7). Of 171 cases, 71% (n = 122) occurred in blacks and 2% (n = 3) occurred in Asian-Pacific islanders. Age 0 to 4 years (107 of 171) compared with age 5 to 14 years (64 of 171) was an additional risk factor for the development of tuberculosis (odds ratio, 3.4; 95% confidence interval 2.5, 4.7)), whereas gender was not. Males accounted for 49% of cases (83 of 171). During the period 1983 to 1993 there was no trend of increasing or decreasing numbers among child cases (trend test P = 0.94) despite significant changes by year. The purified protein derivative test had a 9% (8 of 89) false negative rate and was significantly more likely to be negative in children younger than 1 year (4 of 12 vs. 4 of 77; P = 0.01). During the 2-year interval 1992 to 1993, 19% of cases were thought to be preventable. We believe that the PPD skin test is useful and an improved contact investigation is essential to preventing childhood tuberculosis. Miniepidemics of transmission of tuberculosis from adults to a large group of children partially explain the observed disease pattern.

Identification of a contaminating Mycobacterium tuberculosis strain with a transposition of an IS6110 insertion element resulting in an altered spoligotype.

ABSTRACT Molecular fingerprinting with the IS6110insertion sequence is useful for tracking transmission ofMycobacterium tuberculosis within a population or confirming specimen contamination in the laboratory or through instrumentation. Secondary typing with other molecular methods yields additional information as to the relatedness of strains with similar IS6110 fingerprints. Isolated, relatively rare, random events within the M. tuberculosis genome alter molecular fingerprinting patterns with any of the methods; therefore, strains which are different by two or more typing methods are usually not considered to be closely related. In this report, we describe two strains of M. tuberculosis, obtained from the same bronchoscope 2 days apart, that demonstrated unique molecular fingerprinting patterns by two different typing methods. They were closely linked through the bronchoscope by a traditional epidemiologic investigation. Genetic analysis of the two strains revealed that a single event, the transposition of an IS6110 insertion sequence in one of the strains, accounted for both the differences in the IS6110pattern and the apparent deletion of a spacer in the spoligotype. This finding shows that a single event can change the molecular fingerprint of a strain in two different molecular typing systems, and thus, molecular typing cannot be the only means used to track transmission of this organism through a population. Traditional epidemiologic techniques are a necessary complement to molecular fingerprinting so that radical changes within the fingerprint pattern can be identified.

Preventable Childhood Tuberculosis in Alabama: Implications and Opportunity

Childhood tuberculosis (TB) cases indicate recent community transmission and thus reflect the effectiveness of TB control efforts, particularly the contact investigation. Objective. To evaluate all preventable childhood TB cases and implications in the context of TB morbidity trends. Design. Statewide morbidity trends are presented from 1983 to 1997. Since 1992, each child TB case is classified as either preventable or not preventable, based on a standard definition. Main Outcome Measures. Case characteristics (preventable and not preventable), TB disease rates over time, and reasons for preventable case classification. Setting. Alabama TB control program, from January 1, 1983 through December 31, 1997. Results. For the period 1983–1997, nonwhite children had a higher disease rate (rate ratio: 5.7; 95% confidence interval: 4.3,7.6) than white children. Since 1990, the overall child rate has increased significantly despite a decline in the adult rate. Among 120 child cases diagnosed from 1992 to 1997, 25 (21%) were classified as preventable. The causes were contact investigation interview failure (12/25 = 48%), delay to evaluation (16%), source case noncompliance with previously prescribed preventive therapy (16%), and source case diagnosed out of state (16%) with no initial investigation performed in Alabama. All preventable cases identified were black children; the proportion of preventable cases did not vary by age group or sex. During 1996, the case rate for nonwhite children exceeded that of adult whites. Conclusions. Childhood TB in Alabama for nonwhites is rising despite a national downward trend. TB is clearly a disproportionate disease burden for the states African American population, and the median case age is falling. Additional research and improved training in contact investigation are required to assess this situation and effectively intervene.

Nesidioblastosis Causing Reversal of Insulin-Dependent Diabetes and Development of Hyperinsulinemic Hypoglycemia

The recent publication of a case of non-insulin-dependent diabetes mellitus (NIDDM) reversed by an insulinoma (1) stimulated us to review our records of a patient who was seen by us for a short period of time in 1981. This patient presented with the features of insulin-dependent diabetes mellitus (IDDM) and ketoacidosis. Subsequently, the need for insulin was reversed in spite of a massive weight gain, and later fasting hyperinsulinemic hypoglycemia occurred due to nesidioblastosis.

Interleukin-2 augmentation of interleukin-1 and prostaglandin E2 production.

Some of the major side effects of interleukin‐2 (IL‐2) therapy in the treatment of malignancies may be related to increased interleukin‐1 (IL‐1) and/or prostaglandin E2 (PGE2) production. We examined the effect of recombinant (rIL‐2) on the in vitro production of IL‐1β and PGE2 by unstimulated and LPS‐activated human blood mononuclear cells (PBMC). We also compared the effect of rIL‐2 on IL‐1β production by adherent and nonadherent blood mononuclear cell populations. Cultures of PBMC (5 × 106/ml) were incubated for 24 hr in media only (control), 1,000 U/ml rIL‐2, 2 μg/ml LPS, or both LPS and rIL‐2. Supematants obtained from these cultures were analyzed for levels of IL‐1β and PGE2 by radioimmunoassays. The addition of rIL‐2 caused an increase in IL‐1β production in 13 of 13 control PBMC cultures and in 11 of 13 LPS‐stimulated cultures, which were significant increases as determined by paired f tests. When PBMC were fractionated into plastic adherent and nonadherent populations, the rIL‐2 induced increases in IL‐1β production were more consistent in control (six of seven cases) and LPS (seven of seven cases) cultures of plastic nonadherent cells than in control (three of seven cases) and LPS (four of seven cases) cultures of plastic adherent cells. Recombinant IL‐2 did not increase PGE2 production in control PBMC cultures (none of four cases), but did so in LPS‐stimulated PBMC cultures (three of four cases)). These results suggest that rIL‐2 may increase IL‐1 production in vivo and thus possibly account for some of the side effects of this therapy.