Faculty Bibliography

Human granulocytic ehrlichiosis (HGE) is an emerging and occasionally fatal human infectious disease whose pathogenesis is largely unknown. Goodman et al. (1) recently described the successful cultivation of the HGE infectious agent in human promyelocytic HL-60 leukemic cells. It was reported in the same study that infectivity invariably led to host cell death, although the mechanism by which HGE infection triggers cellular self-destruction is as yet undetermined. In this communication, we show that in vitro passage of HGE pathogen-infected blood elicits a significantly dysfunctional G1-to-S transition. Moreover, we provide evidence that the cytopathic properties of the HGE pathogen are attributed to its ability to induce apoptosis in host HL-60 cells. Determination of specific protein expression changes by Western blot analysis showed that HGE infection resulted in reduced expression of PCNA and pRB, both of which play a role in cell cycling. Moreover, the steady state level of bcl-2, which protects eukaryotic cells against apoptosis, is suppressed by exposure to the HGE agent. These results suggest that this pathogen HGE induces apoptosis in HL-60 cells by a mechanism involving the shut-off of multiple cell cycle and apoptosis regulatory events

In 10 consecutive patients with an acute febrile illness, human granulocytic ehrlichiosis was confirmed with specific polymerase chain reaction studies, serologic conversion, or both. Although no patients had the clinical features most suggestive of early Lyme disease (eg, erythema migrans or cranial nerve palsy), tests for antibody to Borrelia burgdorferi produced a reaction in most patients. In 6 of 7 patients (86%) with evaluable results, enzyme-linked immunosorbent assay yielded positive or equivocal findings, and an immunoblot technique yielded positive findings in 60% to 90% of patients, depending on the criteria used for interpretation. Inasmuch as approximately 25% of nymphal Ixodes scapularis ticks in Westchester County, New York, are infected with B burgdorferi, the probability that at least 9 of these patients were coinfected with B burgdorferi and human granulocytic ehrlichiosis by the same tick bite is estimated to be .00003. These observations suggest that serodiagnosis is insufficient to establish the presence of coinfection with B burgdorferi

BACKGROUND: Human granulocytic ehrlichiosis (HGE) is a newly described illness with few reports in the literature. OBJECTIVE: To describe the clinical and laboratory feature of HGE. DESIGN: Case series. SETTING: Tertiary care facility in New York State. PATIENTS: 18 adult patients with HGE. MEASUREMENTS: Epidemiologic, clinical, and laboratory features; treatment; and outcome of patients with HGE. RESULTS: Patients presented with such symptoms as fever (94%) and myalgia or arthralgia (78%). Thirteen patients (71%) recalled being bitten by a tick before onset of symptoms. Leukopenia or thrombocytopenia was seen in 82% of patients, and abnormal liver enzyme levels were seen in 81%. Results of polymerase chain reaction were positive in 9 of 12 patients (75%); morulae were seen in 3 of 12 patients (25%); and the agent that causes HGE was cultured from 2 patients. All but one patient (94%) developed antibodies to Ehrlichia equi. Five patients (28%) were briefly hospitalized, and none died. All patients were successfully treated with doxycycline. CONCLUSIONS: The illness associated with HGE in these patients from the northeastern United States was more mild than that originally described in reports of HGE in the midwestern United States

OBJECTIVE: The purpose of this study was to evaluate amniotic fluid lactate dehydrogenase level in comparison with other rapid markers in prediction of microbial invasion of the uterine cavity and preterm delivery < or = 36 hours after amniocentesis. STUDY DESIGN: One hundred thirty-one women in preterm labor with intact membranes underwent transabdominal amniocentesis. Amniotic fluid was analyzed for leukocyte count, glucose level, lactate dehydrogenase level, and Gram stain. Cultures for aerobes, anaerobes, and Mycoplasma sp. were performed. Amniocentesis-to-delivery interval was calculated. The study group was divided and the findings compared according to amniotic fluid culture results and according to amniocentesis-to-delivery interval. Sensitivity, specificity, and positive and negative predictive value were calculated for lactate dehydrogenase, leukocyte count, glucose, and Gram stain in the prediction of positive amniotic fluid culture and preterm delivery < or = 36 hours after amniocentesis. Receiver-operator characteristic curve analysis, logistic regression analysis, t tests, and nonparametric tests were used. RESULTS: The prevalence of positive amniotic fluid cultures was 12% (16 of 131). The median lactate dehydrogenase level (1084 U/L) was significantly greater for women with a positive amniotic fluid culture than for those with a negative culture (median lactate dehydrogenase level 194 U/L; p < 0.0002). The critical values calculated for optimal performance in prediction of a positive amniotic fluid culture were a lactate dehydrogenase level > or = 419 U/L, leukocyte count > or = 50 cells/mm3 (50 x 10(6)/L) and glucose < or = 17 mg/dl (0.94 mmol/L). Lactate dehydrogenase, leukocyte count, glucose, and Gram stain were equally sensitive and specific in prediction of a positive amniotic fluid culture. Thirty-nine women (29.8%) gave birth < or = 36 hours after amniocentesis. The median lactate dehydrogenase level (414 U/L) was significantly greater among women giving birth < or = 36 hours after amniocentesis than among women giving birth > 36 hours after amniocentesis (median lactate dehydrogenase, 173 U/L; p < 0.001). Critical values of lactate dehydrogenase > or = 225 U/L, leukocyte count > or = 10 cells/mm3 (10 x 10(6)/L) and glucose < or = 34 mg/dl (1.9 mmol/L) were selected for optimal performance in prediction of amniocentesis-to-delivery interval < or = 36 hours. Lactate dehydrogenase level had the best sensitivity (74%) in prediction of delivery < or = 36 hours after amniocentesis in contrast to leukocyte count (49%), glucose (62%), and positive Gram stain (26%). Amniotic fluid lactate dehydrogenase values > or = 225 U/L were associated with a fivefold greater risk for delivery < or = 36 hours after amniocentesis (odds ratio 5.46, 95% confidence interval 2.00 to 14.87; p = 0.0006). CONCLUSION: Amniotic fluid lactate dehydrogenase level has diagnostic value in prediction of a positive amniotic fluid culture and delivery < or = 36 hours after amniocentesis. Lactate dehydrogenase is a readily available, inexpensive, rapid amniotic fluid marker that can be measured in any hospital laboratory

BACKGROUND: A 2-test approach for the serologic diagnosis of Lyme disease has recently been proposed. A positive or equivocal result on a first-stage test (eg, an enzyme immunoassay) is followed by a Western immunoblot test. For a sample to be considered seropositive for Lyme disease, the immunoblot result must be positive. OBJECTIVES: To assess the accuracy of IgM immunoblotting for detection of early Lyme disease and to establish interpretative criteria for a commercially available immunoblot assay. METHODS: Serum samples from 44 patients with erythema migrans were tested by an IgM immunoblot assay. All patients were culture-positive for Borrelia burgdorferi. Serum samples from 2 different control groups were also tested. Interpretative criteria were developed using receiver operating characteristic curves. RESULTS: The presence of any 2 IgM bands was found to be the optimal criterion for a positive test result, and in patients with illness of less than 7 days' duration, this was significantly more sensitive than the criterion of any 2 of the 3 specific bands defined by the Centers for Disease Control and Prevention/Association of State and Territorial Public Health Laboratory Directors Lyme Disease Workgroup (P < .05). Specificity of the criterion of any 2 bands was 100% for 1 group of controls but only 96% for the more clinically relevant control group; this small difference had a large impact on the positive predictive value in populations at low risk for Lyme disease. CONCLUSIONS: Using a commercially available immunoblot test kit, the presence of any 2 IgM bands is proposed as a positive result. The predictive value of a positive IgM immunoblot result, however, is poor in patients with minimal clinical evidence for Lyme disease

BACKGROUND: The diagnosis of erythema migrans (EM), the characteristic rash of early Lyme borreliosis, is based primarily on its clinical appearance since it often occurs prior to the development of a specific antibody response. Other skin disorders, however, may be confused with EM. METHODS: Between June 1991 and September 1993, a prospective study was conducted at the Lyme Disease Diagnostic Center of the Westchester County Medical Center to isolate Borrelia burgdorferi systematically from patients with Em, and to characterize the clinical manifestations of patients with culture-documented infection. Skin biopsies and/or needle aspirates of the advancing margin of primary lesions, and blood specimens from adult patients were cultured for B burgdorferi in modified Barbour-Stoenner-Kelly medium at 33 degrees C. RESULTS: B burgdorferi was recovered from 79 patients (49 [62%] males) ranging in age from 16 to 76 years old (mean, 43 +/- 14 years old). Maximum EM diameter (mean, 16 +/- 10 cm; range, 6-73 cm) was a function of EM duration (mean 6.7 +/- 6.4 days; range, 1-39 days) (correlation coefficient = 0.7; P < 0.001). Twenty (25%) patients had noted a tick bite at the site of the primary lesion a mean of 10 days (range, 1-27 days) before onset. Multiple EM lesions (range, 2-70) were present in 14 (18%) patients. Systemic symptoms were present at the time of culture in 54 patients (68%) including fatigue (54%), arthralgia (44%), myalgia (44%), headache, (42%), fever and/or chills (39%), stiff neck (35%), and anorexia (26%). Thirty-three patients (42%) had at least one objective finding on physical examination in addition to EM, including 18 (23%) with localized lymphadenopathy, 13 (16%) with fever (t > or = 37.8 degrees C), seven (9%) with tender neck flexion, six (8%) with joint tenderness, and 1 each with joint swelling, nuchal rigidity, and facial nerve palsy. No patient had new electrocardiogram evidence of atrioventricular block. Liver function assays were abnormally elevated in 37% of patients. Thirty-four percent of patients were seropositive by enzyme-linked immunosorbent assay at presentation. Most others rapidly seroconverted so that 69 of 78 evaluable patients (88%) were seropositive at some point during the first month after diagnosis. CONCLUSIONS: We describe the largest group of culture-positive patients with EM from the United States to date. Although systemic symptoms were present in most patients, objective evidence of advanced disease was uncommon. Our patients with culture-confirmed EM were less sick than those described in the days before culture confirmation was possible. The ability to isolate B burgdorferi from lesional skin of large numbers of patients with EM should make culture-positive patients the standard by which to define manifestations of early Lyme borreliosis associated with this rash. Microbiologic documentation of Lyme borreliosis will help delineate the manifestations of this illness, and should form the framework for research directed at pathophysiology, diagnosis, treatment, and prevention

We investigated the appearance and evolution of immunoglobulin M (IgM) and IgG antibodies to Borrelia burgdorferi in 46 patients with culture-proven erythema migrans (EM). All patients received antimicrobial treatment and were prospectively evaluated for up to 1 year. A total of 257 serially collected serum samples were tested by commercial IgG-IgM enzyme-linked immunosorbent assay and separate IgM and IgG immunoblots (IBs). At the baseline, 33% of the patients had a positive ELISA result and 43% of the patients had a positive IgM IB result by using the criteria of the Centers for Disease Control and Prevention-Association of State and Territorial Public Health Laboratory Directors for the interpretation of IB results. Positive serology at the baseline and the rate of seroconversion correlated directly with disease duration and/or evidence of dissemination prior to treatment. At days 8 to 14 after the baseline, 91% of patients had a positive ELISA result and/or IgM IB result. Peak IgM antibody levels were seen at this time in patients with localized or disseminated disease. The most frequent IgM bands at the baseline and the peak were of 24 kDa (OspC), 41 kDa, and 37 kDa. Although 89% of the patients developed IgG antibodies as determined at a follow-up examination, only 22% were positive by the IgG IB criteria of the Centers for Disease Control and Prevention-Association of State and Territorial Public Health Laboratory Directors. The persistence of antibodies was directly related to disease duration and/or dissemination prior to treatment. Since IgM antibodies to the 24- and 41-kDa antigens remained detectable for long periods, 38% of IgM IBs were still positive at 1 year postbaseline. IgM to antigens of 39, 58, 60, 66, or 93 kDa, conversely, were most often seen in sera obtained within 1 month postbaseline. Their presence may be of assistance in confirming a recent infection with B. burgdorferi in individuals living in areas where Lyme disease is endemic

The purpose of our study was to determine whether Borrelia burgdorferi spirochetes were present in placentas of asymptomatic women with reactive Lyme serology using a silver stain, and to confirm the identity of the spirochetes by polymerase chain reaction (PCR). Sixty placentas of asymptomatic women with ELISA-positive or-equivocal serology for Lyme antibodies during pregnancy were examined for spirochetes using a silver stain. The results of the ELISA serology were confirmed by Western blot analysis. PCR amplification for B. burgdorferi was performed on placentas identified to have spirochetes and on a group of placentas negative for spirochetes. Spirochetes were identified by silver staining in 3 (5%) of the 60 placentas. PCR confirmed B. burgdorferi nucleotide sequences in 2 of the placentas. The 5 women had equivocal Lyme ELISA and negative syphilis serology. The results of the Western blot analysis were negative in 2 cases and indeterminate in 1 case. Six controls were negative for spirochetes by silver staining and PCR. A normal perinatal outcome was observed in all cases. Spirochetes identified in placental tissue of pregnancies with reactive Lyme serology were confirmed by PCR to be B. burgdorferi. There was no relationship between the presence of placental spirochetes and the results of Lyme serology or the pregnancy outcome

Using a commercially available enzyme-linked immunosorbent assay (ELISA) and an immunoblot assay (IB), we tested sera from 100 patients with erythema migrans (EM) seen in 1991 a the Westchester County Medical Center Lyme Disease Diagnostic Center. Convalescent-phase sera were available from 59 patients. Fifty-five patients had EM of < 7 days' duration, 31 had EM of 7 to 14 days' duration, and 14 had EM of > 14 days' duration. During the acute phase of infection, 35 patients had a positive ELISA result and 43 had a positive IB result by the recently published criteria of Dressler et al. (F. Dressler, J. A. Whalen, B. N. Reinhardt, and A. C. Steere, J. Infect. Dis. 167:392-400, 1993) for interpretation of IB in patients with Lyme disease. A greater sensitivity of IB was observed in patients with EM of < 7 days' duration, as follows: 14 of 55 (25%) for IB versus 7 of 55 (13%) for ELISA (P = 0.144). Sera of all 14 patients with EM of > 14 days' duration were reactive by both tests, as follows: 13 positive and 1 equivocal by ELISA and 12 positive and 2 indeterminate by the IB. The band reactivity most frequently observed in the IB was to the 41- and 25-kDa antigens, the latter being the most frequent band observed in immunoglobulin M blots. Seroconversion was observed in 74 and 64% of evaluable patients by ELISA and IB, respectively, despite the use of antibiotic therapy