Julia A. Kiehlbauch

Helicobacter cinaedi-associated bacteremia and cellulitis in immunocompromised patients.

Although initially associated with gastroenteritis in homosexual men [1-4], Helicobacter cinaedi was also isolated from asymptomatic [1, 3, 4] and bacteremic [5-7] homosexual men. These organisms are also associated with illness outside the homosexual population; Vandamme and colleagues [8] described three bacteremic patients and two children with fecal isolates. A retrospective epidemiologic study was done to define the clinical spectrum of illness and epidemiologic risk factors associated with H. cinaedi infection in the United States. We also looked at laboratory methods used to recover H. cinaedi. Patients were identified from H. cinaedi isolates received at the Centers for Disease Control and Prevention (CDC) between January 1982 and August 1990. Case Report A 32-year-old man developed red-copper-colored blotches on his left ankle 6 weeks before admission; these spread to his right ankle, up the legs, to the arms, chest, and face. At the time of onset, the patient reported fever but no gastrointestinal symptoms. He was given cephalexin, 500 mg four times a day, for sinusitis; he noted that the rash worsened. Three weeks before he was hospitalized, the patient received oral ciprofloxacin for 2 weeks; the rash resolved. The rash reappeared, and the patient presented with chills and fever (a temperature of 39.4 C), nausea, arthralgias, and the maculopapular skin eruption. At admission, blood cultures were done, and he was treated empirically with cefotaxime and tobramycin. His leukocyte count at admission was 7.8 109/L, with a differential of 73 segmented neutrophils, 18 lymphocytes, and 8 monocytes. Platelets were noted as adequate. His medical history included seropositivity to human immunodeficiency virus (HIV) and transient immune thrombocytopenia (platelet count, 35 109/L); a platelet count of 192 109/L was noted 4 months before admission. The patient stated that the rash first appeared approximately 24 hours after using a whirlpool spa. He did not recall eating raw dairy products, eggs, seafood, or other meats. The patient reported recreational exposure to sea and lake water but did not recall drinking untreated surface water. He traveled within the United States and Europe and reported having homosexual contact in the month before onset of illness. After positive blood cultures were identified, the patient was given empiric ciprofloxacin, 250 mg twice a day for 14 days. The cellulitis cleared, and the patient was discharged. The cellulitis recurred 11 weeks later; the patient was rehospitalized and treated with cefotaxime and ciprofloxacin. Two blood cultures obtained at the time of the second admission again yielded H. cinaedi. The patients leukocyte count was now 6.8 109/L. The rash again cleared. The patient reported three additional recurrences. After zidovudine therapy was initiated, no further recurrence of cellulitis was noted. Results Patients with H. cinaedi infection ranged in age from 24 to 84 years (mean, 44 years); 83% were men. Patients resided in 14 different states (1, Arizona; 1, Colorado; 6, California; 1, Georgia; 1, Illinois; 1, Kansas; 1, Michigan; 2, Missouri; 2, Nebraska; 1, New Mexico; 2, Ohio; 1, Tennessee; 3, Texas; and 1, Wisconsin). Isolation of the organism occurred from 1982 to 1990 (2 in 1982, 2 in 1984, 3 in 1985, 2 in 1986, 3 in 1987, 5 in 1988, 2 in 1989, and 4 in 1990) without seasonal clustering. No one died of this infection. Appendix Table 2 shows the clinical features of 21 patients (information on 5 patients was provided after a review of medical records by the local health department). Most patients had a sudden onset of fever (range, 37.8 to 40.0 C; mean, 38.9 C). Nine bacteremic patients (41%) had both fever and a distinctive cellulitis that was described as atypical, appearing red-brown or copper without noticeable warmth. Underlying immunosuppressive illness was reported for 14 of 21 patients; other underlying conditions, previously associated with systemic Campylobacter infections, were reported infrequently. Appendix Table 1. Characteristics of Patients with Helicobacter cinaedi Infection Information regarding potential exposures in the 4 weeks before onset was available for 15 patients. The most frequent exposures of interest were contact with or consumption of untreated surface water (three patients) and contact with animals (nine patients). Four patients reported out-of-state travel in the 4 weeks before onset: one to Mexico, one to Europe, one to Hawaii, and one to Colorado on a camping trip. All blood isolates of H. cinaedi were recovered after detection by an automated blood culture instrument; 21 of 22 isolates were recovered from the aerobic bottle; 1 isolate was also detected in the anaerobic bottle, and 1 isolate was detected solely in the anaerobic bottle. Most isolates were detected after 5 or more days of incubation by slightly elevated growth indexes (generally between 40 and 80; mean, 57). In general, organisms were not seen on initial Gram staining of the blood culture material but were detected by dark-field or acridine orange staining. Only 9 (41%) of 22 blood isolates were recovered by the primary hospital laboratory; all other isolates were cultured by reference laboratories. In contrast to Campylobacter jejuni, H. cinaedi is not susceptible to erythromycin in vitro (Table 1). In general, tetracyclines and aminoglycosides seem most effective in vitro. Table 1. Antimicrobial Susceptibility of 22 Strains of Helicobacter cinaedi Associated with Human Clinical Illness* Discussion Our findings indicate that H. cinaedi is associated with a new syndrome, consisting of fever, bacteremia, and recurrent cellulitis. Most patients had signs of systemic infection, including leukocytosis, and were often thrombocytopenic. Although H. cinaedi isolates were recovered primarily from immunocompromised patients and from those with chronic alcoholism, we also documented infections in three nonimmunocompromised men and in women (both with and without HIV infection). Thus, the patient group affected by H. cinaedi is larger than originally thought. We did not find distinctive risk factors for acquisition of H. cinaedi by interviewing a subset of patients; however, our review may have been hampered by the time between illness and interview. Our data suggest that contact with animals or exposure to untreated surface water are possible sources of infection. Currently, H. cinaedi has been isolated only from humans and gerbils [9], but no patients reported having contact with gerbils. Many antimicrobial therapies were used to treat patients with H. cinaedi infection. From our series, it appears that treatment with a penicillin, tetracycline, or aminoglycoside may be more effective than treatment with cephalosporins, erythromycin, or ciprofloxacin. In addition, prolonged therapies (2 to 6 weeks) may be more effective than short-term therapies ( 10 days; data not shown). The apparent effectiveness of tetracycline or aminoglycosides agrees with in vitro antimicrobial susceptibility data. Although two reports suggest treating H. cinaedi with ciprofloxacin [10, 11], infection reappeared in two patients treated with this agent, and our in vitro data indicate that their isolates and two additional isolates were resistant (minimal inhibitory concentration > 8 g/mL). This finding suggests that ciprofloxacin should be used with caution. However, we were unable to obtain isolates before and after treatment, so we cannot determine whether resistance to ciprofloxacin developed as a result of therapy or existed before therapy was begun. Our susceptibility data agree with previously published data [12], with one exception: Fifty percent of our strains were resistant to cephalothin. Laboratory diagnosis of H. cinaedi infection is unlikely using blood culture procedures that rely on visual detection because it grows slowly and the growth is difficult to see; all blood isolates were recovered using an automated system. We therefore suggest examining blood cultures that develop slightly elevated growth indexes in an automated system using acridine orange staining, Giemsa staining, or dark-field examination before discarding a specimen as negative. In general, specialized culture techniques and prolonged incubation (7 days) must be used to isolate these organisms: Growth is enhanced by the presence of hydrogen gas in a microaerobic atmosphere and incubation on rich, nonselective media (blood or chocolate agar) at 37 C. Techniques that would probably isolate H. cinaedi from stool specimens include filtration onto nonselective media or inoculation of appropriate selective media and incubation at 37 C in a hydrogen-containing atmosphere for 3 to 4 days. A retrospective review of patients with H. cinaedi infection suggests a syndrome of recurrent febrile bacteremia, which may be accompanied by cellulitis in immunocompromised patients. Helicobacter cinaedi infection should be considered in an immunocompromised or thrombocytopenic patient with fever and cellulitis. Specific antimicrobial therapy may be needed to prevent recurrence. The slow growth and fastidious culture requirements of this organism indicate that it may be currently under-recognized.

The use of plasmid profiles and nucleic acid probes in epidemiologic investigations of foodborne, diarrheal diseases

The application of nucleic acid analyses to investigations of infectious disease outbreaks has resulted in useful molecular strain markers that distinguish the epidemic clone of a particular pathogen and help identify specific vehicles of infection. We have successfully used plasmid profile analysis, restriction endonuclease digestion of plasmid and whole-cell DNAs, and nucleic acid hybridization to investigate recent outbreaks of foodborne diarrheal illness. Plasmid analysis has been important in identifying epidemic strains of Salmonella enteritidis and Escherichia coli O157:H7. In a culture survey of S. enteritidis isolates from humans and a variety of animals, including chickens and chicken eggs, we identified 16 distinct plasmid profiles and used these to differentiate strains, especially within commonly occurring phage types (Colindale 8 and 13a). HindIII digests of plasmid DNA were useful in distinguishing plasmids of similar mass but dissimilar enzyme target sequences; they clearly distinguished S. enteritidis strains causing systemic infections in children in parts of Africa from U.S. isolates. Investigations of outbreaks of hemorrhagic colitis have also been assisted by plasmid analysis. Restriction endonuclease digests of whole-cell DNA and Southern blot analysis, hybridizing with E. coli 16S and 23S rRNA (ribotyping), have been effective subtyping techniques, especially for plasmidless isolates of Campylobacter jejuni. In five outbreaks of C. jejuni infections, ribotyping of PvuII and ClaI digests distinguished individual epidemic strains within one commonly occurring C. jejuni serotype (Penner 2, Lior 4). Preliminary data show that ribotyping of NcoI digests can also distinguish individual epidemic strains of E. coli O157:H7 and may provide a more stable marker than plasmid profiles. Specific DNA probes derived from cloned virulence genes of E. coli have been invaluable in epidemic investigations and surveys. Using colony hybridization, we found in one survey of stool specimens from 174 dairy cattle that 11% of animals were asymptomatically carrying Shiga-like toxigenic E. coli other than O157:H7. We also found that newly synthesized oligonucleotide probes for the Shiga-like toxins I and II agreed 100% with cloned gene probes in a study of 613 E. coli strains. Future studies of these organisms will include the use of additional synthetic oligonucleotides as primers to amplify the toxin genes directly in patient and animal specimens by the polymerase chain reaction. There is a continuing and expanding role for molecular approaches in epidemiological investigations. The DNA methods described above are not based on the often complex expression of phenotypic characteristics, and, unlike sensitive and specific techniques such as phage typing, a single method can be used to study a variety of Gram-positive and negative bacterial pathogens.

In vitro susceptibilities of aerotolerant Campylobacter isolates to 22 antimicrobial agents.

We evaluated the in vitro activities of 22 antimicrobial agents against 78 human and animal isolates belonging to two aerotolerant Campylobacter species, C. cryaerophila and C. butzleri, using a broth microdilution technique. An additional 10 antimicrobial agents were included at concentrations found in selective Campylobacter media. Strains of C. cryaerophila belonged to two DNA hybridization groups: DNA hybridization group 1A, which includes the type strain of C. cryaerophila, and DNA hybridization group 1B. The aminoglycosides, fluoroquinolones, and one tetracycline (minocycline) demonstrated the most activity against all DNA hybridization groups (C. cryaerophila DNA groups 1A and 1B and C. butzleri). Most isolates were resistant to cephalosporin antibiotics, with the exception of cefotaxime, and were variably susceptible to trimethoprim-sulfamethoxazole. C. cryaerophila DNA hybridization group 1A isolates were generally susceptible to the tetracyclines, chloramphenicol, nalidixic acid, azithromycin, erythromycin, and roxithromycin and moderately susceptible to clindamycin, trimethoprim-sulfamethoxazole, ampicillin, and ampicillin-sulbactam. The MICs of tetracyclines were higher for C. butzleri and C. cryaerophila DNA hybridization group 1B isolates than for C. cryaerophila DNA hybridization group 1A isolates, but most strains were still susceptible to doxycycline and tetracycline; all isolates were susceptible to minocycline. C. butzleri and C. cryaerophila DNA hybridization group 1B isolates were generally resistant to the macrolide antibiotics (including erythromycin), chloramphenicol, clindamycin, nalidixic acid, ampicillin, and trimethoprim-sulfamethoxazole. Differences in antimicrobial susceptibility between aerotolerant Campylobacter species and more common Campylobacter species, e.g., C. jejuni, suggest that different treatment strategies may be necessary. Strains of all three DNA hybridization groups of aerotolerant Campylobacter isolates were susceptible to colistin, polymyxin B, and rifampin at concentrations commonly used in selective media. These results suggest that primary isolation methods for Campylobacter species may need to be modified to include aerotolerant Campylobacter strains.

Meningococcal Carriage Among Georgia and Maryland High School Students

BACKGROUND Meningococcal disease incidence in the United States is at an all-time low. In a previous study of Georgia high school students, meningococcal carriage prevalence was 7%. The purpose of this study was to measure the impact of a meningococcal conjugate vaccine on serogroup Y meningococcal carriage and to define the dynamics of carriage in high school students. METHODS This was a prospective cohort study at 8 high schools, 4 each in Maryland and Georgia, during a school year. Students at participating schools received quadrivalent meningococcal conjugate vaccine that uses diphtheria toxoid as the protein carrier (MCV4-DT). In each state, 2 high schools were randomly assigned for MCV4-DT receipt by students at the beginning of the study, and 2 were randomly assigned for MCV4-DT receipt at the end. Oropharyngeal swab cultures for meningococcal carriage were performed 3 times during the school year. RESULTS Among 3311 students, the prevalence of meningococcal carriage was 3.21%-4.01%. Phenotypically nongroupable strains accounted for 88% of carriage isolates. There were only 5 observed acquisitions of serogroup Y strains during the study; therefore, the impact of MCV4-DT on meningococcal carriage could not be determined. CONCLUSIONS Meningococcal carriage rates in US high school students were lower than expected, and the vast majority of strains did not express capsule. These findings may help explain the historically low incidence of meningococcal disease in the United States.

Diversity of factor H-binding protein in Neisseria meningitidis carriage isolates.

Several meningococcal vaccines under development for prevention of serogroup B disease target the factor H-binding protein (FHbp), an immunogenic lipoprotein expressed on the surface of Neisseria meningitidis. Based upon sequence and phylogenetic analyses, FHbp can be classified into 3 protein variants (1, 2 or 3) or 2 subfamilies (A or B). The potential effect of FHbp-containing vaccines on meningococcal carriage is not known. We determined the diversity of FHbp among a population of carriage isolates obtained from Georgia and Maryland high school students in 1998 and 2006-2007. Analysis of the fHbp gene sequence from 408 carriage isolates identified 30 different FHbp protein sequences. The majority of carriage isolates harbored FHbp proteins belonging to variant 2/subfamily A. Association between FHbp proteins and genetic lineage was observed among the carriage isolates. However, split decomposition analysis, together with tests of linkage disequilibrium and pairwise homoplasy suggest recombination at fHbp contribute to allelic diversity. Of note, the FHbp proteins in serogroup B vaccines under development are either absent or not well represented in this carriage population. The FHbp genetic repertoire observed in carriage isolate populations will be useful in understanding the potential impact of FHbp-containing vaccines on meningococcal carriage.