Importance of holotoxin assembly in Ptl-mediated secretion of pertussis toxin from Bordetella pertussis.

We examined the structural components of pertussis toxin that are required for efficient export from Bordetella pertussis via the Ptl system, a member of the type IV family of macromolecular transporters. First, we constructed a strain of B. pertussis that contains a functional Ptl system but does not produce pertussis toxin. Plasmids which express either the S1 subunit or the B oligomer were then introduced into this strain. We found that the B oligomer of the toxin is not secreted in the absence of the S1 subunit. Conversely, the S1 subunit is also not secreted by a Ptl-mediated mechanism in the absence of the B oligomer. Thus, an assembled holotoxin is required for Ptl-mediated export of pertussis toxin from B. pertussis.

Identification and characterization of PtlC, an essential component of the pertussis toxin secretion system.

PtlC is a member of a set of proteins necessary for the secretion of pertussis toxin (PT) from Bordetella pertussis. Using polyclonal antibodies specific for PtlC, we identified PtlC as a protein with an apparent molecular weight of 85,000 that localizes to the membrane fraction of bacterial cell extracts. We found that a mutant strain of B. pertussis that contains an in-frame deletion in ptlC is unable to secrete PT and that PT secretion is fully restored by expressing ptlC in trans, indicating that PtlC is essential for transport of PT across the bacterial membrane(s). PT secretion was inhibited in wild-type B. pertussis after introduction of a plasmid expressing a mutant ptlC altered in the putative nucleotide-binding region, suggesting that this region of PtlC is essential for proper function. Moreover, the observed dominant negative phenotype suggests that PtlC either functions as a multimer or interacts with some other component(s) necessary for secretion of PT.

Evidence for a ninth gene, ptlI, in the locus encoding the pertussis toxin secretion system of Bordetella pertussis and formation of a PtlI-PtlF complex.

The pertussis toxin secretion system of Bordetella pertussis initially was thought to comprise eight proteins, PtlA-PtlH. We have investigated the existence of another protein, PtlI, encoded by a putative gene located between ptlD and ptlE. A B. pertussis strain expressing a ptlI::phoA translational fusion possessed alkaline phosphatase activity, suggesting that ptlI encodes a protein. In B. pertussis, a protein with an apparent molecular weight of approximately 5,200 (similar to that predicted by the ptlI sequence) was immunoreactive with an antibody raised to a PtlI-maltose-binding protein fusion protein. PtlE expression in a mutant sustaining an in-frame deletion in ptlI indicated that ptlE starts further downstream than initially predicted. PtlF, not detected in the ptlI deletion mutant, was restored partially by expressing ptlI in trans. A 36-kDa species, consistent with a PtlI-PtlF complex, was immunoreactive with antibodies to PtlI and PtlF in nonreduced cell extracts of a Bordetella bronchiseptica strain which overexpresses the Ptl proteins. Upon dithiothreitol treatment, the 36-kDa species was diminished greatly or undetectable. In B. pertussis, PtlI and PtlF co-precipitated with antibody to PtlF. These findings demonstrate the existence of PtlI and a PtlI-PtlF complex, providing the first description of an interaction between Ptl proteins.

Distribution of CD4+ T lymphocytes at diagnosis of acquired immunodeficiency syndrome-defining and other human immunodeficiency virus-related illnesses. The Adult and Adolescent Spectrum of HIV Disease Project Group.

BACKGROUND: Depletion of circulating CD4+ T lymphocytes among persons infected with the human immunodeficiency virus (HIV) is associated with increased risk for development of opportunistic, life-threatening diseases and death. METHODS: To describe the levels of CD4+ T lymphocytes at which acquired immunodeficiency syndrome (AIDS)-defining and other illnesses initially occur, we analyzed data from an ongoing survey of medical records of 18,062 HIV-infected patients who received medical care between January 1990 and August 1993 in more than 100 clinics, hospitals, and private practices in 10 US cities. We report the median and upper 80th percentile CD4+ T-lymphocyte counts at diagnosis. RESULTS: We found that AIDS-defining conditions first occurred in HIV-infected patients with CD4+ T-lymphocyte counts below 0.20 x 10(9)/L (200/microL) for 80% of diagnoses. Similarly, AIDS-defining diseases occurred at counts below 0.05 x 10(9)/L for 50% of diagnoses. Exceptions to both criteria were invasive cervical cancer and pulmonary tuberculosis. Non-AIDS-defining illnesses with which 80% of patients were diagnosed at CD4+ T-lymphocyte counts below 0.20 x 10(9)/L were bacterial sepsis and retinopathy (excluding cytomegalovirus). CONCLUSION: Our observations support the need for continued CD4+ cell count monitoring below a level of 0.20 x 10(9)/L as a guide to diagnosis and medical management of HIV-infected persons.

Pneumococcal and influenza vaccination levels among HIV-infected adolescents and adults receiving medical care in the United States. Adult and Adolescent Spectrum of HIV Disease Project Group.

OBJECTIVE: To assess pneumococcal and influenza vaccination coverage among HIV-infected adolescents and adults receiving medical care in the United States. DESIGN: Periodic medical record reviews. SETTING: More than 90 clinics, hospitals, and private medical practices in nine cities. PATIENTS: HIV-infected individuals aged > or = 13 years were included in the analyses of pneumococcal (n = 9737) and influenza (n = 6161) vaccination coverage. MAIN OUTCOME MEASURES: Documentation of receipt of pneumococcal and influenza vaccines in medical records during 6-18-month and 12-month periods, respectively. RESULTS: Overall, 37 and 33% of individuals received pneumococcal and influenza vaccines, respectively. In general, vaccination levels varied little by age group, race/ethnicity, or mode of HIV exposure. Having had at least five medical visits was significantly associated with having received pneumococcal and influenza vaccines [adjusted odds ratio (OR), 1.7 for each]. Having a CD4+ T-lymphocyte count < 200 x 10(6)/l (adjusted OR, 0.8) and being female (adjusted OR, 0.7) were associated with non-receipt of pneumococcal vaccine. Lower pneumococcal vaccination coverage among women was mostly accounted for by pregnancy. CONCLUSION: Until new, more effective means of preventing pneumococcal disease and influenza become available, efforts should be directed towards improving vaccination levels among HIV-infected individuals.

Measles vaccination in pediatric emergency departments during a measles outbreak.

OBJECTIVE: To determine the proportion of preschool-aged patients attending two inner-city hospital pediatric emergency departments (EDs) who were eligible for measles vaccination, to describe their demographic and clinical characteristics, and to assess the performance of the ED immunization programs that were implemented during a measles outbreak in vaccinating eligible children. DESIGN: Cross-sectional study. SETTING: Pediatric EDs of two urban hospitals in Chicago, Ill, in 1989. PARTICIPANTS: Children 6 months to 5 years of age seen in the EDs. INTERVENTION: None. MAIN OUTCOME MEASURES: The proportion of preschool-aged patients attending the two EDs who were eligible for measles vaccination and the proportion of vaccine-eligible children who were given measles vaccine. RESULTS: Of 508 ED patients at hospital A and 255 patients at hospital B, 18% and 29%, respectively, were considered to be vaccine eligible. The most common discharge diagnoses of eligible patients were viral syndrome, otitis media, and minor trauma. Of vaccine-eligible patients, 59% at hospitals A and B were not vaccinated in the ED. At hospital B, patients with an infectious or respiratory disease diagnosis were less likely to be vaccinated than those with other diagnoses (P < .05). CONCLUSIONS: Many children seen in these EDs were eligible for measles vaccination, and many eligible patients were not vaccinated. During community outbreaks of measles, optimal vaccination programs in pediatric EDs could increase vaccination coverage among inner-city preschool-aged children who may have limited access to health care.

Pertussis in Missouri: evaluation of nasopharyngeal culture, direct fluorescent antibody testing, and clinical case definitions in the diagnosis of pertussis.

No diagnostic test for pertussis in routine use in the United States has both high sensitivity and high specificity. During a statewide increase in the incidence of pertussis in Missouri, we studied the clinical features of 153 patients with suspected pertussis in the Greater St. Louis area from whom a specimen for pertussis culture had been taken between 15 May and 19 September 1989. In this cross-sectional study, nasopharyngeal cultures were more likely to be positive for persons whose specimens were collected < 21 days after cough onset (adjusted rate ratio [RRa] and 95% confidence interval = 3.4; 1.5-8.0) and who were not receiving erythromycin/sulfamethoxazole prior to the culture [RRa = 5.8; 0.8-40.6], who had received fewer than three prior doses of pertussis vaccine [RRa = 1.8; 0.8-4.2], and whose specimen was in transit to the laboratory for < 4 days [RRa = 2.0; 0.8-5.5]. Among children < 5 years of age, spasmodic cough plus a lymphocytosis of > 10,000/mm3 was the acute symptom complex associated with the highest predictive value for a positive culture result (67%). Cough for > or = 14 days plus whoop was sensitive (81%) and specific (58%) for identifying children with culture-confirmed pertussis. Direct fluorescent antibody staining performed well as a screening test for pertussis but requires substantial commitment of personnel and resources. In the absence of a positive culture result, clinical case definitions should be used for decision making (e.g., initiation of antimicrobial therapy and routine case reporting).

Fatal respiratory disease due to Corynebacterium diphtheriae: case report and review of guidelines for management, investigation, and control.

Dramatic reductions in the incidence of diphtheria and high levels of childhood vaccination in recent decades have led the United States to establish the goal of diphtheria elimination among persons < or = 25 years of age by the year 2000. In 1990, an unimmunized 25-month-old child died of respiratory diphtheria in Dade County, Florida, before treatment with diphtheria antitoxin could be instituted. Twenty-three asymptomatic household contacts and other close contacts of the child were identified, cultured for Corynebacterium diphtheriae, given antimicrobial prophylaxis, and vaccinated with diphtheria toxoid when indicated. Three contacts (13%) had pharyngeal cultures positive for toxigenic C. diphtheriae of the same type as that causing infection in the deceased child, but no additional cases developed. Although the source of infection was not determined, three other close contacts had recently been to Haiti, where diphtheria is endemic. A serological survey of 396 children < 5 years of age who received care at a medical center in Dade County revealed that 22% lacked protective immunity to diphtheria. Attainment of the goal of diphtheria elimination among persons < or = 25 years of age--and ultimately among all persons--will depend on the maintenance of a high level of clinical awareness of the disease, the prompt institution of preventive measures among close contacts of patients with sporadic cases, and improved vaccination levels among infants, children, and adults.

Projections of the number of persons diagnosed with AIDS and the number of immunosuppressed HIV-infected persons--United States, 1992-1994.

This report presents projections of the number of persons who will initially be diagnosed with a condition included in the 1987 surveillance definition for acquired immunodeficiency syndrome (AIDS) in the United States during the period 1992-1994. The report also presents estimates and projections of the prevalence of persons infected with the human immunodeficiency virus (HIV) who have CD4+ T-lymphocyte (T-cell) counts < 200/microL and who have not been diagnosed with a condition listed in the 1987 AIDS surveillance definition. These estimates and projections are used to predict the effect of expanding the AIDS surveillance definition to include all HIV-infected persons with a CD4+ T-cell count < 200/microL. Approximately 58,000 persons were diagnosed with AIDS in the United States during 1991. During the period 1992-1994, the number of persons newly diagnosed with AIDS is expected to increase by at most a few percent annually, with approximately 60,000-70,000 persons diagnosed per year. Although AIDS diagnoses among homosexual and bisexual men and among injecting drug users are projected to reach a plateau during this period, the number of AIDS diagnoses among persons whose HIV infection is attributed to heterosexual transmission of HIV is likely to continue to increase through 1994. The number of living persons who have been diagnosed with AIDS is expected to increase from approximately 90,000 in January 1992 to approximately 120,000 in January 1995. There is, however, considerable uncertainty in these projections. For example, the plausible range for the number of persons initially diagnosed with AIDS in 1994 is 43,000-93,000. CDC estimates that, as of January 1992, 115,000-170,000 U.S. residents had severe immunosuppression (a CD4+ T-cell count < 200 cells/microL without a diagnosis of AIDS in an HIV-infected person). Only about 50,000 of these persons were receiving medical care for HIV-related conditions and were known to have a CD4+ T-cell count < 200 cells/microL. The number of persons with severe immunosuppression is expected to increase to 130,000-205,000 by January 1995, with the actual number more likely to be in the lower half of this range than the upper half. The expanded AIDS surveillance definition, which includes severe immunosuppression, is predicted to result in an increase of approximately 75% in the number of persons reported during 1993, but an increase of < 20% in 1994 compared with the number of persons who would have been reported had the definition not been changed.(ABSTRACT TRUNCATED AT 400 WORDS)

Vaccination levels and missed opportunities for measles vaccination: a record audit in a public pediatric clinic.

A record audit of 254 children attending a public clinic in Los Angeles was conducted to assess immunization levels prior to a measles outbreak in the community. Coverage with all vaccines appropriate for age decreased from 67% at 3 months to 25% at 19 months. Delay in initiating vaccination was associated with increasing risk for delayed measles-mumps-rubella vaccine beyond age 2 years (P less than .05). In one third of children, health care providers missed an opportunity to administer measles-mumps-rubella vaccine. Recall systems and elimination of missed opportunities may increase vaccination levels in clinic populations. Record audits should be considered for use in guiding the management of immunization programs.

Spectrum of disease in persons with human immunodeficiency virus infection in the United States.

OBJECTIVE: To describe the spectrum of disease in persons with human immunodeficiency virus (HIV) infection. DESIGN: Retrospective survey of medical records. SETTING: More than 50 clinics, hospitals, and private medical practices in nine US cities. PATIENTS: A total of 626 women and 7008 men 13 years of age or older with HIV infection who received medical care from January 1990 through March 1991 were consecutively enrolled. MAIN OUTCOME MEASURES: Any history of diseases in the 1987 case definition for the acquired immunodeficiency syndrome (AIDS), and during the 12-month period preceding enrollment (baseline period), the occurrence of other major diseases, hospitalizations, and results of CD4+ lymphocyte counts. RESULTS: Thirty-two percent of persons met the 1987 case definition for AIDS. The occurrence of an AIDS-indicator disease during the baseline period ranged from 3% (33/1011) to 46% (1254/2748) among persons with CD4+ lymphocyte counts of 0.50 x 10(9)/L or greater and fewer than 0.20 x 10(9)/L (greater than or equal to 500 and less than 200 CD4+ lymphocytes per microliter), respectively, and, at comparable CD4+ lymphocyte levels, was similar among women compared with men, and among persons who reported intravenous drug use compared with men who reported male-to-male sex. The frequency of one or more other major infectious diseases (eg, other pneumonias, bacterial sepsis, pulmonary tuberculosis) ranged from 6% to 16% among persons with CD4+ lymphocyte counts of 0.50 x 10(9)/L or greater and fewer than 0.20 x 10(9)/L, respectively; these illnesses were also associated with a history of intravenous drug use. Among persons who did not meet the 1987 AIDS case definition, 30% of those with an available CD4+ lymphocyte count had fewer CD4+ cells than 0.20 x 10(9)/L, 8% had one or more major infectious diseases, and 14% had one or more hospital admissions. CONCLUSIONS: For every person with AIDS at these sites, two additional persons with HIV infection were receiving medical care, many of whom had severe immunosuppression and a broad spectrum of serious HIV-related disease.

Epidemiological features of pertussis in the United States, 1980-1989.

From 1980 through 1989, 27,826 cases of pertussis were reported to the Centers for Disease Control, for an average annual crude incidence of 1.2 cases/100,000 population. The incidence of reported disease increased in all age groups during this period, but the increase was disproportionately large among adolescents and adults. Infants between 1 and 2 months of age were at highest risk for pertussis (average annual incidence, 62.8/100,000). Infants less than 2 months of age had the highest reported rates of pertussis-associated hospitalization (82%), pneumonia (25%), seizures (4%), encephalopathy (1%), and death (1%). Rates of complication were generally higher among unvaccinated children than among those who had received three or more doses of diphtheria-tetanus-pertussis vaccine; 64% of children 3 months to 4 years of age who had reported cases of pertussis had not been immunized appropriately for their age. Whereas control of pertussis in the United States may be further improved through increased levels of diphtheria-tetanus-pertussis vaccination among eligible infants and children, the use of acellular vaccines in adolescents and adults may also be needed to reduce the burden of pertussis in very young infants.

Pediatric emergency room visits: a risk factor for acquiring measles.

In recent years, measles outbreaks have occurred among unimmunized children in inner cities in the United States. From May 1988 through June 1989, 1214 measles cases were reported in Los Angeles, and from October 1988 through June 1989, 1730 cases were reported in Houston. More than half of cases were in children younger than 5 years of age, most of whom were unvaccinated. Of cases of measles in preschool-aged children, nearly one fourth in Los Angeles and more than one third in Houston were reported by one inner-city emergency room. To evaluate whether emergency room visits were a risk factor for acquiring measles, in Los Angeles, 35 measles patients and 109 control patients with illnesses other than measles, and in Houston, 49 measles patients and 128 control patients, who visited these emergency rooms, were enrolled in case-control studies. Control patients were matched to case patients for ethnicity, age, and week of visit. Records were reviewed to determine whether case patients had visited the emergency room during the period of potential measles exposure, which was defined as 10 to 18 days before rash onset, and whether control patients had visited 10 to 18 days before their enrollment visit. In Los Angeles, 23% of case patients and 5% of control patients (odds ratio = 5.2, 95% confidence interval = 1.7, 15.9; P less than .01), and in Houston, 41% of case patients and 6% of control patients (odds ratio = 8.4, 95% confidence interval = 3.3, 21.2; P less than .01), visited the emergency room during these periods.(ABSTRACT TRUNCATED AT 250 WORDS)