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Ticks introduce a variety of pharmacologically active molecules into their host during attachment and feeding in order to obtain a blood meal. People who are repeatedly exposed to ticks may develop an immune response to tick salivary proteins. Despite this response, people usually are unaware of having been bitten, especially if they are not repeatedly exposed to ticks. In order to develop a laboratory marker of tick exposure that would be useful in understanding the epidemiology of tick-borne infection and the immune response to tick bite, we developed an enzyme-linked immunosorbent assay (ELISA) to detect antibody to a recombinant form of calreticulin protein found in the salivary glands of Ixodes scapularis , a member of a complex of Ixodes ticks that serve as the vectors for Lyme disease, human babesiosis, and human granulocytic anaplasmosis. Using this assay, we tested sera obtained from C3H/HeN and BALB/c mice before and after experimental deer tick infestation. These mice developed antibody to Ixodes calreticulin antigen after infestation. We then used the same assay to test sera obtained from people before and after they experienced deer tick bite(s). People experiencing deer tick bite(s) developed Ixodes calreticulin-specific antibody responses that persisted for up to 17 months. This Ixodes recombinant calreticulin ELISA provides objective evidence of deer tick exposure in people.

Background The importance of vascular occlusion in the pathogenesis of human haemoprotozoal disease is unresolved. Methods Giemsa-stained tissue sections from a human case of Babesia microti infection in a splenectomized patient with chronic lymphocytic leukaemia and colon cancer were examined to ascertain the distribution of parasitized erythrocytes within the vascular lumen. Results No evidence of sequestration was observed. Conclusion This first report on the vascular location of B. microti in human tissue suggests that severe multi-organ failure due to babesiosis is independent of sequestration of parasitized erythrocytes. A similar pathogenesis may also cause multi-organ failure in other intraerythrocytic protozoal infections, including falciparum malaria.

A newborn infant whose condition was diagnosed as herpes simplex encephalitis and who had subsequent recurrences of skin disease had repeated episodes of neutropenia while receiving therapy with intravenous (30 mg/[kg.d]) or oral (30 mg/]kg.d]) acyclovir. The neutropenia did not recur when the dosage of oral acyclovir was reduced to 10 mg/(kg.d). This case represents the first well-documented report of acyclovir-induced neutropenia.

Human granulocytic ehrlichiosis (HGE) is an emerging tick-borne infectious disease caused by Anaplasma phagocytophilum. Clinical features include a flu-like illness that usually resolves within 1 week. More serious infection may occur that requires hospital admission or culminates in death. Doxycycline is the treatment of choice for HGE but may cause permanent staining of teeth in children younger than 8 years of age. We report successful treatment of HGE with rifampin in 2 children, 4 and 6 years old. A course of rifampin for 5 to 7 days should be considered in children younger than 8 years of age who experience non-life-threatening A phagocytophilum infection.

Human infection due to Babesia microti has been regarded as infrequent and a condition primarily affecting the elderly or immunocompromised. To determine whether risk in endemic sites may be increasing relative to that of Borrelia burgdorferi and to define its age-related clinical spectrum, we carried out a 10-year community-based serosurvey and case finding study on Block Island, Rhode Island. Less intensive observations were conducted in nearby sites. Incidence of babesial infection on Block Island increased during the early 1990s, reaching a level about three-fourths that of borrelial infection. The sera of approximately one-tenth of Block Island residents reacted against babesial antigen, a seroprevalence similar to those on Prudence Island and in southeastern Connecticut. Although the number and duration of babesial symptoms in people older than 50 years of age approximated those in people 20 to 49 years of age, more older adults were admitted to hospital than younger adults. Few Babesia-infected children were hospitalized. Babesial incidence at endemic sites in southern New England appears to have risen during the 1990s to a level approaching that due to borreliosis.

Preface. Stealth Parasites: The Under Appreciated Burden of Parasitic Zoonoses in North America J.L. Gauthier, et al. Toxocariasis and Baylisascariasis P.M. Schantz. Trichinellosis D.D. Despommier. Larval Tapeworm Infections: Cysticercosis, Cystic Echinococcosis and Alveolar Echinococcosis A. Flisser. Intestinal Tapeworm Infections D.J. Richardson. Other Noteworthy Zoonotic Helminths D.J. Richardson. Cryptosporidiosis C. Chappell. Toxoplasmosis D.S. Lindsay. Babesiosis P.J. Krause. Other Noteworthy Zoonotic Protozoa O.J. Pung. Zoonotic Arthropod Parasites L.A. Durden. Index.

Objective The objective of this quality standard is to optimize the treatment of bacteremia in hospitalized patients by ensuring that the antibiotic given is appropriate in terms of the blood culture susceptibility of the pathogen. Although this standard may appear to be minimal in scope, it is needed because appropriate antimicrobial treatment is not given in 5%-17% of cases. To implement the standard, physicians, pharmacists, and microbiologists will need to devise a coordinated strategy. Options We considered criteria for appropriate dosing, most cost-effective selection, proper antibiotic levels in serum, least toxicity, narrowest spectrum, specific clinical indications, and optimal duration of treatment. All these criteria were rejected as the basis for the standard because they were too controversial and too difficult to be applied by a nonphysician chart reviewer. In contrast, the selection of an antibiotic to which the pathogen is sensitive is a noncontroversial criterion that is easy for a chart reviewer to apply. Outcomes The standard is designed to reduce the incidence of adverse outcomes of septicemia, such as renal failure, prolonged hospitalization, and death. Evidence Several well-designed clinical trials without randomization as well as case-controlled studies have confirmed the benefit of using an antibiotic that is appropriate in light of the susceptibility of the isolate in blood culture. Prospective, randomized, placebo-controlled trials are not available. Values Our premise is that the presence of bacteremia is a risk factor for serious adverse outcomes. We also believe that the administration of antibiotics must always be guided by the susceptibility report for the pathogen(s) obtained from blood cultures. This concern is more critical for pathogens from the blood than for those from most other body sites. We had evidence that susceptibility reports for pathogens from positive blood cultures were not always used properly. We used group discussion to reach a consensus among the members of the Quality Standards Subcommittee. Benefits, harms, and costs Through the implementation of this standard, at least 5% of cases of bacteremia could be treated more appropriately. An unknown number of deaths would likely be prevented, and mortality from bacteremia treated inappropriately would probably be reduced. The primary undesirable feature of the standard is an increased workload for pharmacists and microbiologists. Recommendations Treatment of bacteremia with an antibiotic that is appropriate in terms of the pathogen's blood-culture susceptibility is a minimal standard of care for all patients. Validation We consulted more than 50 experts in infectious diseases from the fields of medicine, surgery, pediatrics, obstetrics and gynecology, nursing, epidemiology, pharmacology, and government. In addition, the methods for its implementation were reviewed by the American Society of Hospital Pharmacists and were tested by one of the members of the Quality Standards Subcommittee. Sponsors The Quality Standards Subcommittee of the Clinical Affairs Committee of the Infectious Diseases Society of America (IDSA) developed the standard. The subcommittee was composed of representatives of the IDSA (P.A.G. and J.E.M.), the Society for Hospital Epidemiology of America (R.P.W.), the Surgical Infection Society (E.P.D.), the Pediatric Infectious Diseases Society (P.J.K.), the Centers for Disease Control and Prevention (W.J.M.), the Obstetrics and Gynecology Infectious Diseases Society (R.L.S.), and the Association of Practitioners of Infection Control (T.L.B.). Funding was provided by the IDSA and the other cooperating organizations. The standard is endorsed by the IDSA.

Epidemiological methods are needed to evaluate community exposure to Borrelia burgdorferi, the causative agent of Lyme disease (LD). For LD serodiagnosis, the Centers for Disease Control and Prevention (CDC) recommends a 2-test approach that involves enzyme immunoassay (EIA) testing and Western immunoblotting (WB) of EIA-equivocal and EIA-positive specimens. The specificity of this approach was evaluated among residents of a LD-endemic community and was compared with WB alone and with a simplified 2-test approach (WB of equivocal EIA only). Participants reporting no previous diagnosis of LD were recruited during a community-wide serosurvey on Block Island, Rhode Island. Of 80 eligible participants, 20 had received LD vaccine. Seven (35%) of 20 vaccinees and 22 (37%) of 60 nonvaccinees reported nonspecific symptoms compatible with LD in the previous year. In this highly LD-endemic community, the overall specificity of the CDC-recommended approach was highest (100%), followed by WB alone (98.7%), then the simplified approach (95%).

Objective. A recombinant lipoprotein outer surface protein A (OspA) Lyme disease (LD) vaccine (LYMErix) has been shown to be safe and effective in preventing LD in adults and in adolescents 15 years of age and older. Children are at risk for developing LD. This clinical study was conducted to address the safety and immunogenicity of LD vaccine in children 4 to 18 years of age. Methods. A randomized, placebo-controlled clinical trial was conducted at 17 investigational sites in Lyme-endemic areas in the United States. Immunogenicity data from this study also were compared with data obtained from the adult efficacy study. A total of 4090 healthy children and adolescents (age range: 4–18; mean age: 10.4 years) were randomized; 4087 were vaccinated, and a subset of 301 children participated in the immunogenicity analysis. Children were randomized to receive either 30 μg of LD vaccine (N = 3063) or placebo (N = 1024) on a 0, 1, 12-month schedule. Safety assessments evaluated both solicited (local: redness, swelling, and pain; general: fever, headache, fatigue, arthralgia, and rash) and unsolicited adverse events. Serum specimens were collected at month 0 or month 2, and months 6, 12, and 13. Results. Solicited reactogenicity data revealed a higher incidence of local injection site reactions and general symptoms (fever, headache, fatigue, and arthralgia) in vaccine than placebo recipients. The majority of events were limited in duration (mean: 2–3 days) and were mild to moderate in severity. The total IgG anti-OspA geometric mean titer (GMT) in the pediatric vaccine recipients at month 13 was as good as and statistically higher than the GMT in the adult cohort at month 13 (27 485 enzyme-linked immunosorbent assay units [EL.U]/mL vs 8216 EL.U /mL). All of the pediatric vaccine recipients attained a level of antibody concentration ≥1400 EL.U/mL (proposed seroprotective level) compared with 90% of adults attaining levels ≥1400 EL.U/mL in the efficacy trial. Conclusions. LD vaccine administered on a 0, 1, 12-month schedule generally is well tolerated and immunogenic in children 4 to 18 years of age. The safety profile consists of mild to moderate local injection site reactions and flu-like symptoms of limited duration and did not worsen with subsequent injections. IgG GMT at month 13 was threefold higher than the month 13 GMT obtained in the adult efficacy study. This higher immune response in children should provide protection against LD.

SUMMARY Babesiosis is an emerging, tick-transmitted, zoonotic disease caused by hematotropic parasites of the genus Babesia. Babesial parasites (and those of the closely related genus Theileria) are some of the most ubiquitous and widespread blood parasites in the world, second only to the trypanosomes, and consequently have considerable worldwide economic, medical, and veterinary impact. The parasites are intraerythrocytic and are commonly called piroplasms due to the pear-shaped forms found within infected red blood cells. The piroplasms are transmitted by ixodid ticks and are capable of infecting a wide variety of vertebrate hosts which are competent in maintaining the transmission cycle. Studies involving animal hosts other than humans have contributed significantly to our understanding of the disease process, including possible pathogenic mechanisms of the parasite and immunological responses of the host. To date, there are several species of Babesia that can infect humans, Babesia microti being the most prevalent. Infections with Babesia species generally follow regional distributions; cases in the United States are caused primarily by B. microti, whereas cases in Europe are usually caused by Babesia divergens. The spectrum of disease manifestation is broad, ranging from a silent infection to a fulminant, malaria-like disease, resulting in severe hemolysis and occasionally in death. Recent advances have resulted in the development of several diagnostic tests which have increased the level of sensitivity in detection, thereby facilitating diagnosis, expediting appropriate patient management, and resulting in a more accurate epidemiological description.

Increased recognition of the prevalence of human babesiosis in the United States, together with rising concern about the potential for transmission of this infection by blood transfusion, has provided motivation to develop definitive serologic and molecular tests for the causative agent, Babesia microti . To develop more sensitive and specific assays for B. microti , we screened a genomic expression library with patient serum pools. This screening resulted in the identification of three classes of novel genes and an additional two novel, unrelated genes, which together encode a total of 17 unique B. microti antigens. The first class (BMN1-2 family) of genes encodes seven closely related antigens with a degenerate six-amino-acid repeat that shows limited homology to Plasmodium sp. merozoite and sporozoite surface antigens. A second class (BMN1-8 family) of genes encodes six related antigens, and the third class (BMN1-17 family) of genes encodes two related antigens. The two remaining genes code for novel and unrelated sequences. Among the three classes of antigens and remaining novel sequences, five were chosen to code for the most immunodominant antigens (BMN1-2, -9, -15, and -17 and MN-10). Western blot analysis with the resulting recombinant proteins indicated that these antigens were targets of humoral immune responses during B. microti infection in humans.

Human babesiosis in the United States is caused predominantly by Babesia microti , a tick-transmitted blood parasite. Improved testing methods for the detection of infection with this parasite are needed, since asymptomatic B. microti infection represents a potential threat to the blood supply in areas where B. microti is endemic. We performed immunoscreening of an expression library of genomic DNA from a human isolate of B. microti (strain MN1). Among 17 unique immunoreactive clones, we identified 9 which represent a related family of genes with little sequence homology to other known sequences but with an architecture resembling that of several surface proteins of Plasmodium . Within this family, a tandem array of a degenerate six-amino-acid repeat (SEAGGP, SEAGWP, SGTGWP, SGTVGP) was found in various lengths between relatively well conserved segments at the N and C termini. In order to examine within-clone variation, we developed a PCR protocol for direct recovery of a specific bmn1-6 homologue directly from 30 human blood isolates, 4 corresponding hamster isolates, and 5 geographically corresponding Peromyscus leucopus (white-footed mouse) isolates. Isolates from the hamsters had the same sequences as those found in the corresponding human blood, suggesting that genetic variation of bmn1-6 does not occur during passage. However, clones from different patients were often substantially different from each other with regard to the number and location of the degenerate repeats within the bmn1-6 homologue. Moreover, we found that strains that were closely related geographically were also closely related at the sequence level; nine patients, all from Nantucket Island, Mass., harbored clones that were indistinguishable from each other but that were distinct from those found in other northeastern or upper midwestern strains. We conclude that considerable genetic and antigenic diversity exists among isolates of B. microti from the United States and that geographic clustering of subtypes may exist. The nature of the bmn1-6 gene family suggests a mechanism of antigenic variation in B. microti that may occur by recombination, differential expression, or a combination of both mechanisms.

The Centers for Disease Control and Prevention (CDC) recommend a two-test approach for the serodiagnosis of Lyme disease (LD), with EIA testing followed by Western immunoblotting (WB) of EIA-equivocal and -positive specimens. This approach was compared with a simplified two-test approach (WB of EIA equivocals only) and WB alone for early LD. Case-patients with erythema migrans (EM) rash >/=5 cm were recruited from three primary-care practices in LD-endemic areas to provide acute- (S1) and convalescent-phase serum specimens (S2). The simplified approach had the highest sensitivity when either S1 or S2 samples were tested, nearly doubling when S2 were tested, while decreasing slightly for the other two approaches. Accordingly, the simplified approach had the lowest negative likelihood ratio for either S1 or S2. For early LD with EM, the simplified approach performed well and was less costly than the other testing approaches since less WB is required.

Babesiosis, a zoonosis caused by the protozoan Babesia microti, is usually not treated when the symptoms are mild, because the parasitemia appears to be transient. However, the microscopical methods used to diagnose this infection are insensitive, and few infected people have been followed longitudinally. We compared the duration of parasitemia in people who had received specific antibabesial therapy with that in silently infected people who had not been treated.Forty-six babesia-infected subjects were identified from 1991 through 1996 in a prospective, community-based study designed to detect episodes of illness and of seroconversion among the residents of southeastern Connecticut and Block Island, Rhode Island. Subjects with acute babesial illness were monitored every 3 months for up to 27 months by means of thin blood smears, Bab. microti polymerase-chain-reaction assays, serologic tests, and questionnaires.Babesial DNA persisted in the blood for a mean of 82 days in 24 infected subjects without specific symptoms who received no specific therapy. Babesial DNA persisted for 16 days in 22 acutely ill subjects who received clindamycin and quinine therapy (P=0.03), of whom 9 had side effects from the treatment. Among the subjects who did not receive specific therapy, symptoms of babesiosis persisted for a mean of 114 days in five subjects with babesial DNA present for 3 or more months and for only 15 days in seven others in whom the DNA was detectable for less than 3 months (P0.05); one subject had recrudescent disease after two years.When left untreated, silent babesial infection may persist for months or even years. Although treatment with clindamycin and quinine reduces the duration of parasitemia, infection may still persist and recrudesce and side effects are common. Improved treatments are needed.

To evaluate the efficacy of an immunoglobulin M indirect immunofluorescent-antibody procedure for diagnosing acute babesiosis, we tested patients with acute babesiosis from a site in New England where the disease is enzootic. The sensitivity of the test was 91%, the specificity was 99%, the positive predictive value was 86%, and the negative predictive value was 99%. This B. microti immunoglobulin M indirect immunofluorescent-antibody procedure is sufficiently sensitive, specific, and reproducible for use in the routine clinical diagnosis of acute babesiosis.

To the Editor: Six weeks after being bitten by a deer tick, a pregnant 31-year-old woman began to have fever, chills, fatigue, and neck pain. During the fifth day of fever, she became jaundiced and was hospitalized. On physical examination, she was 37 weeks pregnant, febrile (temperature, 39.4°C [103°F]), and slightly jaundiced. There was no rash. Laboratory studies showed a hematocrit of 29 percent, a white-cell count of 5400 per cubic millimeter, a platelet count of 96,000 per cubic millimeter, and total bilirubin concentration of 5.4 mg per deciliter. Babesia microti were present in 2 percent of her erythrocytes ( . . .

Introduction Babesiosis is an emerging infectious disease in the Northeast and upper Midwest of the United States. The etiologic agent is Babesia microti, a protozoan parasite that invades red blood cells. In healthy individuals, B. microti infection is typically asymptomatic or evokes a mild flu-like illness that quickly resolves. In immunocompromised patients, particularly those infected with HIV and those treated with rituximab for B cell lymphoma, babesiosis is severe, persistent, and sometimes fatal. To understand the mechanisms by which the host controls and clears B. microti infection, we have developed a mouse model of human babesiosis. Methods Wild-type C57BL/6 mice and C57BL/6 mice that carried a single or a double gene deficiency were infected with B. microti parasitized red blood cells. Parasitemia was assessed by flow cytometry and is defined as the frequency of red blood cells that stain positive for DNA. Babesia specific IgG subclasses were quantified using Babesia infected red blood cells as a source of parasite antigens. Frequencies of interleukin (IL)-21 and/or interferon-gamma (IFNγ) producing T cell subsets were assessed by flow cytometry. Results In wild-type mice, parasitemia was modest and quickly resolved. Lack of mature B cells due to igh6 deficiency did not hinder resistance whereas depletion of CD4+ T cells conferred susceptibility and prevented resolution of infection. Intense, persistent parasitemia was also observed in athymic mice, in rag1−/− mice, and in mice lacking TCRβ or MHC class II. Surprisingly, in cd4−/− mice, parasitemia was intense but resolved. In their spleens, CD8+ T cells and CD4−CD8− (double negative, DN) T cells expanded. Parasitemia resolved in cd4−/− cd8−/− mice, but persisted in cd4−/− igh6−/− mice. Likewise, parasitemia persisted in cd4−/− mice depleted of B cells by repeated administration of anti-CD20. In cd4−/− mice, the rise in circulating Babesia specific IgGs was delayed but concomitant with resolution of parasitemia. Despite the lack of CD4, all subisotypes were produced. IgG1 levels were higher in cd4−/− mice than in wild-type mice whereas IgG3 levels were lower. In cd4−/− mice, DN T cells producing both IL-21 and IFNγ expanded, but not as fast as CD4+ T cells producing both cytokines in wild-type mice. In contrast, DN T cells producing IFNγ alone expanded as fast as their CD4+ T cell counterparts. Furthermore, DN T cells failed to produce IL-21 alone whereas CD4+ T cells producing IL-21 alone quickly expanded. Parasitemia resolved in cd4−/− mice treated with a neutralizing antibody against mouse IFNγ, but persisted in cd4−/− mice that also lacked the IL-21 receptor gene. Despite the requirement for B cells and the production of IgGs, Fcγ receptors were dispensable for resolution of parasitemia in cd4−/− mice. Conclusion Our studies establish that CD4+ T cells are critical for resistance to B. microti infection in C57BL/6 mice and suggest that in the absence of CD4, CD4−CD8− T cells are central for resolution of B. microti parasitemia. In cd4−/− mice, resolution of parasitemia requires B cells and IL-21 but not IFNγ. Given that IgGs are produced but Fcγ receptors are dispensable for host resistance in cd4−/− mice, the role of complement is being investigated.

To determine the risk of acquiring Lyme disease or babesiosis from blood transfusion, serum was collected before and 6 weeks after patients received multiple transfusions during cardiothoracic surgery and antibodies to Borrelia burgdorferi and Babesia microti were measured. Of 155 subjects, 149 received 601 total units of packed red blood cells (PRBC) and 48 received 371 total units of platelets. No patient developed clinical or serologic evidence of Lyme disease; 1 (who received 5 units of PRBC) developed clinical and serologic evidence of babesiosis. The risk of acquiring Lyme disease from a transfused unit of PRBC was 0 (95% confidence interval [CI], 0-0.5%) and from a transfused unit of platelets was 0 (95% CI, 0-0.8%); the same risks for babesiosis were 0.17% (95% CI, 0.004%-0.9%) and 0 (95% CI, 0-0.8%), respectively. The risk of acquiring either Lyme disease or babesiosis from a blood transfusion in Connecticut is very low.

Although tuberculous meningitis is rarely encountered in the United States, clinicians need to have a high index of suspicion for this disease. Intramuscular streptomycin is usually administered as part of a four-drug antituberculous regimen. However, we describe an 8–month-old girl who was diagnosed with Mycobacterium tuberculosis meningitis and received streptomycin intravenously. This route was chosen to avoid daily intramuscular injections because the infant had poor lean muscle mass. The patient's regimen consisted of isoniazid 15 mg/kg/day rifampin 20 mg/kg/day and pyrazinamide 40 mg/kg/day by nasogastric tube, and intravenous streptomycin 15 mg/kg twice/day administered using a controlled-rate infusion pump. The M. tuberculosis strain was subsequently found to be susceptible to all four antituberculous drugs. Her condition improved, and no drug toxicities were observed during her treatment course; isoniazid and rifampin were continued after discharge. The patient was readmitted 1 month later for mental status changes and right-sided weakness. Magnetic resonance scan of the brain revealed numerous solid and ring-enhancing hypointense tuberculomas in the suprasellar cistern, left medial temporal lobe, and brainstem, with significant secondary vasogenic edema as the cause of her symptoms. Although treatment failure was not suspected, cerebrospinal fluid and gastric cultures were tested; all were negative for M. tuberculosis. Dexamethasone was started for treatment of the focalized cerebral edema, presumably occurring from the breakdown of existing tuberculomas, and the patient rapidly improved. She was discharged and continued to receive oral antituberculous therapy for a total of 12 months. At her 1–year follow-up visit, the patient had recovered fully and had no apparent neurologic, otologic, or developmental deficits. The safe and effective use of intravenous streptomycin in this infant suggests that this route of administration may be an alternative to intramuscular streptomycin.

The ability of the neonate to mount an adequate polymorphonuclear leukocyte (PMN) response, either quantitatively or functionally, is impaired. To assess whether neonatal PMN number and function are altered by labor and delivery, three groups of infants were studied: cesarean section without labor (10), cesarean section after labor (10), and vaginal delivery (11). PMN counts were higher in the groups undergoing labor (p less than 0.01) compared with the cesarean section without labor group. Similarly, the labor groups had evidence of complement activation (increased C3a desarg) compared with the cesarean section without labor group. No differences were noted between the groups in measures of PMN motility (chemokinesis or chemotaxis) or PMN degranulation (plasma lysozyme), suggesting that normal labor and delivery does not contribute to the general PMN dysfunction of the neonate.

We found that 4-β-phorbol 12-myristate 13-acetate (PMA) caused decreased expression of the polymorphonuclear neutrophil (PMN) surface antigen 31D8. In contrast to the rapid initiation of the oxidative burst caused by PMA, the effect was slow to start but increased during incubation periods up to 50 min. To study this apparent protein kinase C-independent late effect of PMA, we measured 31D8 expression in PMNs after incubation with various concentrations of PMA. The maximum PMA-induced inhibition was 76 ± 2%, with an ID50 of 3.9 ± 0.4 ng/ml. Oxidants and prooxidants (hydrogen peroxide, hypochlorite, taurine-chloramine, and ferrous iron, with or without H2O2) had no direct effect on 31D8 antigen expression. The following substances were not protective against the inhibitory affect of PMA: (1) antioxidants (superoxide dismutase, catalase, azide, dimethyl sulfoxide, Desferal, and ascorbate, with the exception of α-tocopherol), (2) inhibitors of protein kinase C (H7 and W7), (3) inhibitors of 5-lipoxygenase (A-63162, MK886, and high-dose indomethacin) and (4) inhibitors of cyclooxygenase (low-dose indomethacin). Myeloperoxidase-deficient PMNs had normal 31D8 antigen expression and a decrease of 31D8 antigen expression by PMA, as did normal PMNs. The inactive analog of PMA, 4-α-phorbol didecanoate, had no effect on 31D8 antigen expression. α-Tocopherol (50 μg/ml) and betamethasone (150 μg/ml) protected against the PMA effect by 30.5 ± 7.3 (P < .0005) and 52 ± 15 (P < 0.004) channels, respectively. These results indicate that PMA has a protein kinase C-independent late effect on human neutrophils, which can be prevented by pretreatment with α-tocopherol or the steroid betamethasone. These compounds probably exert their protective effect by membrane stabilization.

We examined the effect of adult fresh frozen plasma (FFP) on neonatal neutrophil (PMN) motility (chemotaxis) using a micropore filter assay. Adult FFP was transfused into 13 neonates receiving FFP transfusion for suspected life-threatening sepsis. Blood was obtained from neonates before and after FFP transfusion for assessment of PMN chemotaxis. An increase in PMN chemotaxis was noted in 12 of the 13 neonates following FFP transfusion, with a mean percentage increase of 12 +/- 3% (p less than 0.01). PMN chemotaxis increased 13 +/- 2% (p less than 0.01) in four bacteremic infants and 11 +/- 5% (p = 0.06) in nine infants without bacteremia. Adult FFP transfusion may enhance impaired neonatal PMN motility and improve outcome from infection in newborn infants.

Immunomodulating agents are being investigated for treatment of infection in newborn infants where morbidity and mortality remain high despite the continued development of new antibiotics. We studied the effect of the methylxanthine pentoxifylline on polymorphonuclear leukocyte (PMN) chemotaxis, F-actin content, and phagocytic activity as measured by nitroblue tetrazolium reduction and H2O2 production in neonates and adults to determine whether pentoxifylline might be useful in augmenting PMN function. The drug was found to have a dose-dependent effect on both neonatal and adult PMN function with enhancement at lower concentrations and suppression at higher concentrations. PMN chemotaxis increased 42% (p less than 0.01) in neonates and 16% (p less than 0.05) in adults at 100 micrograms/mL of pentoxifylline and it decreased 4 and 25%, respectively, at 4000 micrograms/mL. PMN nitroblue tetrazolium reduction increased by 34% in neonates and 23% (p less than 0.05) in adults at 100 micrograms/mL of pentoxifylline and decreased by 52 (p less than 0.01) and 74% (p less than 0.01), respectively, at 2000 micrograms/mL. Similar dose-dependent responses were noted with F-actin content and H2O2 production. These and other observations support the hypothesis that pentoxifylline has a broad range of effects on PMN but that a primary effect is alteration of PMN deformability. Pentoxifylline has potential clinical use as an immunomodulator in augmenting impaired PMN function in neonates and other immunocompromised hosts or in suppressing excessive PMN activity in certain disease processes.

Pentoxifylline (PTX), a drug that improves neutrophil function in vitro, has been shown to protect neonatal mice against death from experimental staphylococcal infection in vivo at a dose of 50 mg/kg. Using a total of 774 neonatal mice, the effects of various doses of PTX were examined and compared with the effects of three analogs: HWA-448, HWA-285, and A81-3138. A subcutaneous abscess was induced with 10(8) Staphylococcus aureus, and drug or saline was given daily subcutaneously from 2 days before to 4 days after infection. Noninfected animals (given saline without S. aureus) had 0% mortality (0 of 66), and infected animals without drug (given saline) had a mortality of 70% (161 of 231). PTX and HWA-448 showed the greatest protection among the drugs tested at 15 mg/kg with mortality rates of 27 and 38%, respectively (Kaplan-Meier method, P = 0.0001 and 0.0004, respectively). HWA-285 was most protective at 25 mg/kg (mortality, 45%; P = 0.0046) and A81-3138 was most protective in animals at 15 mg/kg (mortality, 42%; P = 0.0045). PTX, HWA-448, HWA-285, and A81-3138 at doses of 200, 100, 100, and 50 to 75 mg/kg, respectively, were toxic as shown by worsened weight loss and increased mortality in animals when compared with infected animals without drug. PTX and its analogs decrease mortality from experimental infections at lower doses but are toxic at higher doses. Pharmacokinetic characteristics of the drugs were similar except that HWA-285 produced lower concentrations in serum and A81-3138 showed a dose-dependent kinetics (longer half-life at a higher dose).

Babesiosis is a malaria-like illness caused by the intraerythrocytic parasite Babesia microti and transmitted by the same tick that transmits the Lyme disease spirochete. Both pathogens may be cotransmitted and are endemic in southern New England and New York, the northern Midwest and the far West. The epidemiology and clinical presentation of babesiosis and babesiosis/Lyme disease coinfection are well described in adults but not in children. Accordingly, we reviewed data from longitudinal, clinic-based serosurvey and case-finding studies of tick-borne illness in Connecticut, Massachusets and Rhode Island from 1990 through 1996. A total of 147 episodes of babesial infection were identified, 125 in adults and 22 in children. Approximately a third of children and adults with babesial infection reported symptoms. The most frequently reported symptoms in children were fever (25%), fatigue (19%) and headache (19%) and in adults, fatigue (20%), fever (17%) and myalgias (16%). Children and adults had a similar number of acute symptoms; the average number of symptoms were 4.0 and 4.8, respectively. Children recovered more rapidly, however, as none were ill for more than a month compared with 20% of adults. Coinfection with babesiosis and Lyme disease resulted in more prolonged illness; 17% of children and 26% of adults reported symptoms that lasted for more than a month. We conclude that the frequency of babesial infection is similar in children and adults but children have a shorter duration of illness. Both children and adults experience a more prolonged illness when babesiosis is accompanied by Lyme disease.

Objective. —To determine whether patients coinfected with Lyme disease and babesiosis in sites where both diseases are zoonotic experience a greater number of symptoms for a longer period of time than those with either infection alone. Design. —Community-based yearly serosurvey and clinic-based cohort study. Setting. —Island community in Rhode Island and 2 Connecticut medical clinics from 1990 to 1994. Study Participants. —Long-term residents of the island community and patients seeking treatment at the clinics. Main Outcome Measures. —Seroreactivity to the agents of Lyme disease and babesiosis and number and duration of symptoms. Results. —Of 1156 serosurvey subjects, 97 (8.4%) were seroreactive against Lyme disease spirochete antigen, of whom 14(14%) also were seroreactive against babesial antigen. Of 240 patients diagnosed with Lyme disease, 26 (11%) were coinfected with babesiosis. Coinfected patients experienced fatigue ( P =.002), headache ( P P P =.03), anorexia ( P =.04), emotional lability ( P =.02), nausea ( P =.004), conjunctivitis ( P =.04), and splenomegaly ( P =.01) more frequently than those with Lyme disease alone. Thirteen (50%) of 26 coinfected patients were symptomatic for 3 months or longer compared with 7 (4%) of the 184 patients with Lyme disease alone from whom follow-up data were available ( P P =.06). Conclusions. —Approximately 10% of patients with Lyme disease in southern New England are coinfected with babesiosis in sites where both diseases are zoonotic. The number of symptoms and duration of illness in patients with concurrent Lyme disease and babesiosis are greater than in patients with either infection alone. In areas where both Lyme disease and babesiosis have been reported, the possibility of concomitant babesial infection should be considered when moderate to severe Lyme disease has been diagnosed. ( JAMA . 1996;275:1657-1660)

Since the causative agents of human babesiosis (B) and Lyme disease (LD) are maintained by the same vector ticks in North American sites, human coinfection may occur. To determine whether coinfection is responsible for an increased severity of disease in children, as we have previously found in adults, we compared the clinical manifestations of LD and B alone with that of coinfection in residents of Block Island, RI and southeastern CT. Subjects completed a questionnaire and submitted blood for serological and PCR testing during five successive spring and autumn serosurveys and when they developed symptoms of LD or B from 1990 through 1994. Nineteen percent of the 224 children whose sera reacted against spirochete antigen also were seroreactive against babesial antigen. Of 33 children diagnosed with LD, 21 percent were coinfected. Coinfected children suffered fever, myalgia, chills, headache and cough (P

To assess the possibility of standardization of a commonly used indirect immunofluorescent antibody (IFA) test for detection of Babesia microti antibody in human sera, the results from four reference laboratories were compared. Patients with babesiosis from southern New England (n = 25) and subjects with no history of babesiosis from southern New England (n = 55) and Iceland (n = 50) were enrolled in the study. Anti-Babesia antibody titers were determined in a blinded fashion by IFA test. The range of test results in the four laboratories was 88%-96% sensitivity, 90%-100% specificity, 69%-100% positive predictive value, and 96%-99% negative predictive value. Interlaboratory and intralaboratory concordance ranged from 84% to 85% and 94% to 100%, respectively. This B. microti IFA procedure is a sensitive, specific, and reproducible method for diagnosing babesiosis and is suitable for use as a standard in laboratories testing human sera for B. microti antibody.

Penetration into the middle ear of four antibiotics commonly used in treatment of otitis media was studied by administering a single oral dose of amoxicillin, cefaclor, erythromycin-sulfisoxazole, or trimethoprim-sulfamethoxazole to 83 children with chronic serous otitis media. The antibiotic was given 15-240 min before the removal of middle ear fluid (MEF) by ventilation tubes inserted through the tympanic membrane. At the time MEF was obtained, a sample of blood was drawn from the patient, and concentrations of antibiotic in both specimens were assayed either microbiologically by a disk diffusion method or by high-pressure liquid chromatography. Amoxicillin had the highest ratio of mean peak concentration in MEF to minimal inhibitory concentration (MIC) for the three most common pathogens of otitis media (Streptococcus pneumoniae, ampicillin-sensitive Haemophilus influenzae, and Streptococcus pyogenes), whereas trimethoprim-sulfamethoxazole had the highest ratio of mean peak concentration in MEF to MIC for ampicillin-resistant Haemophilus influenzae.

Newborn infants are at increased risk of morbidity and mortality from infection despite the continued development of new antibiotics. Because impairment of such host defense mechanisms as PMN function is thought to be largely responsible for this problem, correction of these defects in neonates offers a new and potentially important therapy against infection. Further studies are necessary to determine whether transfusion of either adult PMNs, antibody or fresh frozen plasma; administration of immunomodulating drugs; or some combination of these will provide maximum therapeutic benefit for the newborn infant with infection.

Pregnancy is associated with impairment in several immunologic functions that effect the outcome of certain infectious and autoimmune diseases. Previous studies have indicated that pregnant women have decreased neutrophil motility, but results are discordant. We performed a longitudinal study of polymorphonuclear leukocyte chemotaxis using a micropore filter assay and polymorphonuclear leukocyte adherence using a nylon wool column assay in 58 women during pregnancy, delivery, and the postpartum period. Polymorphonuclear leukocyte chemotaxis was decreased significantly in pregnant women during the second (-20%) and third (-17%) trimesters and 1 to 3 months post partum (-18%), but not during the first trimester (-6%) compared with nonpregnant control subjects. The impairment in polymorphonuclear leukocyte chemotaxis appeared to be a cell-associated rather than a humoral defect. Polymorphonuclear leukocyte adherence was decreased significantly in pregnant women during the third trimester (-16%). These data may help explain the increased incidence of infection and amelioration of certain immunologically mediated illnesses during pregnancy.

Polymorphonuclear neutrophil (PMN) adherence to glass and nylon wool was examined in 19 healthy term newborns, 19 normal children aged 2 weeks to 17 years, and 33 adults in good health who were taking no medication. Using a whole blood glass adherence assay, neonatal PMN adherence (0.18 ± 0.16%) was significantly less than PMN adherence in children (7.4 ± 4.0%, P < .0001) and in adults (16.0 ± 4.5%, P < .0001). Similar results were obtained when separated neonatal and adult PMNs were used with both glass and nylon wool column adherence assays, indicating that decreased neonatal PMN adherence is due to a cellular defect. In other experiments there was significantly greater reduction in adherence of separated PMNs with addition of neonatal serum than with adult serum, indicating that a humoral inhibitor also contributes to decreased adherence of neonatal PMNs. Decreased neonatal PMN adherence may be one cause of the increased susceptibility of neonates to serious bacterial infections.

Defects in polymorphonuclear neutrophil (PMN) adherence and chemotaxis in neonates are thought to be an important cause of their increased susceptibility to overwhelming bacterial infection. Few studies of these functions have been carried out in stressed neonates who are at even greater risk of infection. PMN adherence and chemotaxis were examined in 33 stressed neonates with acute lower respiratory illness, 13 healthy neonates, and 43 healthy adults using whole blood PMN adherence and chemotaxis assays. PMN chemotaxis was significantly decreased in stressed neonates (locomotion index of 38.4 +/- 9.7 micron) compared with that of healthy neonates (48.9 +/- 12.8 micron, p less than 0.01) or adults (61.6 +/- 11.9 micron, p less than 0.001). PMN chemotaxis was studied during illness and recovery in 13 of the 33 stressed neonates and showed significant improvement during recovery (41.6 +/- 9.9 and 53.2 +/- 11.9 micron, respectively, p = 0.012). PMN adherence was decreased in stressed neonates (1.4 +/- 1.6%) compared with that of adults (12.3 +/- 11.4%, p less than 0.01) but was similar to that of healthy neonates (1.1 +/- 1.4%). These findings suggest that further impairment of PMN chemotaxis in stressed neonates helps account for their increased susceptibility to overwhelming bacterial infection.

Le test ELISA au rotazyme n'est pas fiable chez les nouveau-nes exclusivement; seulement 7% des examens positifs au rotazyme se sont averes aussi positifs aux tests de confirmation. On preconise de confirmer la positivite des selles ou rotazyme chez les nouveau-nes asymptomatiques ou ceux avec des reactions faiblement positives, a l'aide de la microscopie electronique d'un ELISA de confirmation ou de l'hybridation de l'ARN

We have used a mouse monoclonal antibody (31D8) to determine whether differences in neutrophil (PMN) subpopulations might help explain decreased PMN chemotaxis in neonates compared with that of adults. 31D8 has been shown to bind heterogeneously to adult PMNs. Approximately 80% of the PMNs that strongly bind 31D8 (31D8 “bright”) are the same cells that depolarize and migrate chemotactically when stimulated with the chemoattractant N-formyl-methionylleucylphenylalanine, while the 20% that weakly bind 31D8 fail to similarly respond. All neonatal PMNs bound 31D8 heterogeneously. There was a smaller population of 31D8 “bright” cells in neonates at birth (76% +/- 6%, n = 45) compared with that of neonates at three to 15 days of age (82% +/- 5%, n = 10, P less than 0.002) and both were smaller than that of adults (88% +/- 4%, n = 45, P less than 0.001 and P less than 0.001). Neonatal cord PMNs, which traversed a micropore filter in a modified Boyden chemotaxis chamber in the presence of a chemoattractant, had an increased percentage of 31D8 “bright” cells (89% +/- 7%) than did PMNs which remained above the filter (82% +/- 7%, n = 10, P = 0.034). PMN chemotaxis was less in neonates at birth (32.7 +/- 4.5 micron) than at three to six days of age (36.8 +/- 11.3 micron) and both were decreased compared with that of adults (69.1 +/- 12.4 micron, P less than 0.001 and P less than 0.001). These findings indicate that decreased PMN chemotaxis in neonates may be due in part to a smaller PMN subpopulation of highly motile cells.

Clinical and serologic responses to inactivated influenza virus vaccines were studied in 444 children in Los Angeles under doubld-blind, placebo-controlled protocols. One- and two-dose regimens employing monovalent (A/New Jersey/76) and bivalent (A/New Jersey/76-A/Victoria/75) vaccine were studied in separate trials. In a single dose, whole-virus vaccines were more potent than split-product vaccines as primary inducers of immunity against influenza A/New Jersey/76 virus. Split-product vaccines were better tolerated clinically and, after two-dose regiments, produced rates of seroconversion (greater than 90%) against A/New Jersey/76 virus comparable to those produced by two doses of whole-virus vaccines. Analysis of heterotypic antibody responses induced by monovalent A/New Jersey/76 vaccines revealed only slight increases in antibody against influenza A/Victoria/75 virus. After vaccination with bivalent vaccines, levels of antibody against influenza A/Victoria/75 virus were significantly higher in subjects who received high doses of split-product vaccines than in subjects who received whole-virus vaccines.

Intravenous fat (Intralipid) is used extensively as a major component of parenteral nutrition for patients in the neonatal intensive care unit. Abnormalities of polymorphonuclear leukocyte (PMN) and platelet number or function related to Intralipid infusion have been reported although conflicting results exist. In order to examine potential adverse hematologic effects of Intralipid, 10 ill neonates were studied before and after a 16-hr infusion of 1 g/kilo of Intralipid. PMN count, chemokinesis, chemotaxis, and aggregation were unchanged pre- and post intralipid infusion. Platelet count, bleeding time, and platelet aggregation were also unchanged. Similar results were obtained in vitro when neonatal and adult PMNs and platelets were incubated in Intralipid and their function analyzed. These findings suggest that short-term, low-dose Intralipid has no measurable impact on neonatal PMN or platelet activity and support its use in neonates even in the presence of infection or thrombocytopenia. (Journal of Par...

The pathogenesis of diarrhea following antibiotic therapy for otitis media in young children remains unknown. We performed a prospective study evaluating the incidence of diarrhea and Clostridium difficile toxin in 115 outpatients (ages 6 months to 6 years) with acute otitis media treated with ampicillin, amoxicillin or trimethoprim-sulfamethoxazole. In 21 patients younger than one year of age six of 11 developing diarrhea had toxin-positive stools compared with three of 10 without diarrhea (P = 0.39). In 94 patients between 13 months and 6 years of age three of 12 with diarrhea had toxin-positive stools compared with five of 82 without diarrhea (P = 0.06). Diarrhea was self-limited in all cases. Although the data suggest that C. difficile might have been associated with diarrhea in the older children, further studies will be required to confirm this finding.

Antenatal maternal glucocorticoid administration has been widely used to accelerate fetal lung maturation. Glucocorticoids have also been used postnatally in selected neonates as antiinflammatory agents. Numerous studies have shown that glucocorticoids inhibit multiple components of the immune system including neutrophil (PMN) function in children and adults. Since PMNs are of critical importance in host defense against bacterial infection, impaired PMN function in newborn infants is thought to be an important cause of their increased morbidity and mortality from bacterial infection. Further compromise of neonatal PMN function by exogenous factors such as glucocorticoids may therefore be of significant clinical importance. A micropore filter chemotactic assay was used to determine the in vitro effect of betamethasone on the random migration and directed migration (chemotaxis) of PMNs from 18 neonates. The addition of a concentration of betamethasone (0.01 microgram/ml) similar to that found in cord blood following a standard dose administered to the mother resulted in a significant (p less than 0.01) inhibition in mean neonatal PMN random migration (-15.0 +/- 0.8%) and chemotaxis (-23.5 +/- 3.0%). A similar inhibition was not found when PMNs from 14 adults were exposed to the same concentrations of betamethasone. Betamethasone administration to pregnant women or their newborn infants may further impair PMN motility and lead to an increased morbidity and mortality from bacterial infection in neonates.

During a measles immunization campaign 203 children were enrolled in an antibody response study. Of this group, follow-up clinical data and sera samples were available from 125 children three weeks after immunization and from 90 children ten months later. Seventy-six of the children had been previously vaccinated, ten had a history of measles and 39 denied vaccination or illness. Twenty-six of the children had prevaccination hemagglutination inhibiting antibody titers of less than 5. Of this group 12 had a primary immune response (IgM measles antibody) with geometric mean titers (GMT) of 90 and 40 three weeks and ten months respectively after vaccination. In contrast, the other 14 children with initial titers of less than 5 had secondary immune responses (only IgG measles antibody) with GMTs of 28 and 9 three weeks and ten months after vaccination. Since the antibody responses in these children who had previously been stimulated by measles antigen were modest and transient, it is suggested that booster immunization may not be effective in preventing future secondary vaccine failures. Also noted in this study was a poor correlation between historical data and actual measles antibody.

A new method of micropore filter assay of neutrophil migration requiring only 0.1 ml of whole blood is described and compared with the standard separated polymorphonuclear neutrophil micropore filter assay. Whole blood was added to the upper compartment of a modified Boyden chemotactic chamber, and the neutrophils were allowed to migrate into a cellulose nitrate micropore filter. Acetic acid was used to remove erythrocytes and hemoglobin from the filter. Neutrophil chemotaxis was performed with cells from 19 neonates and 34 adults. The mean neonatal PMN migration was 40% of the adult value (48.1 +/- 12.6 vs 96.8 +/- 16.8 micron) with the whole blood assay and 39% of the adult value (44.7 +/- 10.1 vs 72.9 +/- 22.2 micron) with the separated polymorphonuclear neutrophil assay. The whole blood micropore filter assay, a simple and reliable method of determining neutrophil migration, is especially useful in studying patients when there are difficulties in obtaining large blood samples, such as with neonates and young children.