Horizontal transfer of tetracycline resistance among Chlamydia spp. in vitro.

There are no examples of stable tetracycline resistance in clinical strains of Chlamydia trachomatis. However, the swine pathogen Chlamydia suis is commonly tetracycline resistant, both in America and in Europe. In tested U.S. strains, this resistance is mediated by a genomic island carrying a tet(C) allele. In the present study, the ability of C. suis to mobilize tet(C) into other chlamydial species was examined. Differently antibiotic resistant strains of C. suis, C. trachomatis, and Chlamydia muridarum were used in coculture experiments to select for multiply antibiotic resistant progeny. Coinfection of mammalian cells with a naturally occurring tetracycline-resistant strain of C. suis and a C. muridarum or C. trachomatis strain containing selected mutations encoding rifampin (rifampicin) or ofloxacin resistance readily produced doubly resistant recombinant clones that demonstrated the acquisition of tetracycline resistance. The resistance phenotype in the progeny from a C. trachomatis L2/ofl(R)-C. suis R19/tet(R) cross resulted from integration of a 40-kb fragment into a single ribosomal operon of a recipient, leading to a merodiploid structure containing three rRNA operons. In contrast, a cross between C. suis R19/tet(R) and C. muridarum MoPn/ofl(R) led to a classical double-crossover event transferring 99 kb of DNA from C. suis R19/tet(R) into C. muridarum MoPn/ofl(R). Tetracycline resistance was also transferred to recent clinical strains of C. trachomatis. Successful crosses were not obtained when a rifampin-resistant Chlamydophila caviae strain was used as a recipient for crosses with C. suis or C. trachomatis. These findings provide a platform for further exploration of the biology of horizontal gene transfer in Chlamydia while bringing to light potential public health concerns generated by the possibility of acquisition of tetracycline resistance by human chlamydial pathogens.

Prospective cohort study of microbial and inflammatory events immediately preceding Escherichia coli recurrent urinary tract infection in women.

BACKGROUND: A prospective cohort study was conducted to characterize the temporal sequence of microbial and inflammatory events immediately preceding Escherichia coli recurrent urinary tract infection (rUTI). METHODS: Women with acute cystitis and a history of UTI within the previous year self-collected periurethral and urine samples daily and recorded measurements of urine leukocyte esterase, symptoms, and sexual intercourse daily for 3 months. rUTI strains were characterized by pulsed-field gel electrophoresis and genomic virulence profiling. Urinary cytokine levels were measured. RESULTS: There were 38 E. coli rUTIs in 29 of 104 women. The prevalence of periurethral rUTI strain carriage increased from 46% to 90% during the 14 days immediately preceding rUTI, with similar increases in same-strain bacteriuria (from 7% to 69%), leukocyte esterase (from 31% to 64%), and symptoms (from 3% to 43%), most notably 2-3 days before rUTI (P<.05 for all comparisons). Intercourse with periurethral carriage of the rUTI strain also increased before rUTI (P=.008). Recurrent UTIs preceded by bacteriuria, pyuria, and symptoms were caused by strains less likely to have P fimbriae than other rUTI strains (P=.002). CONCLUSIONS: Among women with frequent rUTIs, the prevalences of periurethral rUTI strain carriage, bacteriuria, pyuria, and intercourse dramatically increase over the days preceding rUTI. A better understanding of the pathogenesis of rUTI will lead to better prevention strategies.

Chlamydia trachomatis laboratory strains versus recent clinical isolates: implications for routine microbicide testing.

A topical microbicide that women can use to prevent sexually transmitted diseases (STDs) is essential, and many microbicide candidates are being tested for activity against human immunodeficiency virus and other STDs, including Chlamydia trachomatis. Screening assays for assessing the activity of microbicides against C. trachomatis are typically done with laboratory-adapted strains, but it is possible that recent clinical isolates may have different susceptibilities to microbicides, as has been seen with Neisseria gonorrhoeae and Lactobacillus spp. (B. J. Moncla and S. L. Hillier, Sex. Transm. Dis. 32:491-494, 2005). We utilized three types of microbicides to help define this aspect of our assay to test microbicides against C. trachomatis in vitro. To simulate conditions of transmission, we used an assay that we previously developed in which we exposed chlamydial elementary bodies to microbicides prior to contact with epithelial cells. We first determined the toxicity of microbicides to the cells used to culture Chlamydia trachomatis in the assay and, if necessary, modified the assay to eliminate toxicity at the concentrations tested. We compared the sensitivities of recent clinical isolates of Chlamydia trachomatis versus laboratory strains of the same serovar and found major differences in sensitivity to nonoxynol-9 (non-9), but only minor differences were seen with the other microbicides. We thus conclude that when assessing activity of potential topical microbicides versus the obligate intracellular bacteria C. trachomatis, the use of recent clinical isolates may not be necessary to draw a conclusion about a microbicide's effectiveness. However, it is important to keep in mind that differences (like those seen with non-9) are possible and that clinical isolates could be included in later stages of testing.

Urinary tract infections in women with type 1 diabetes mellitus: survey of female participants in the epidemiology of diabetes interventions and complications study cohort.

PURPOSE: We determined the prevalence of and risk factors for urinary tract infection in women with type 1 diabetes, and compared the prevalence of cystitis to that in nondiabetic women. MATERIALS AND METHODS: Women enrolled in the Epidemiology of Diabetes Interventions and Complications study were surveyed at year 10 as part of the Uro-EDIC study to assess the prevalence of cystitis and pyelonephritis in the preceding 12 months. Multivariate logistic regression models including measures of glycemic control and vascular complications of type 1 diabetes were used for risk factor analyses. The prevalence of cystitis in Uro-EDIC women was compared to that in a nondiabetic subset of women participants in the National Health and Nutrition Examination Survey III (NHANES III). RESULTS: A total of 550 women participated in the Uro-EDIC survey. The prevalence of cystitis and pyelonephritis in the preceding 12 months was 15% and 3%, respectively. Duration of diabetes, hemoglobin A1C, retinopathy, neuropathy, nephropathy, composite vascular complication score and intensive glycemic therapy during the Diabetes Control and Complications Trial, and Diabetes Control and Complications Trial cohort were not associated with cystitis or pyelonephritis. Sexual activity was associated with increased cystitis risk (adjusted OR 8.28; 95% CI 1.45, 158.32; p = 0.01). The adjusted prevalence of cystitis was 19.1% in Uro-EDIC women and 23.1% in NHANES III participants (adjusted OR 0.78; 95% CI 0.51, 1.22; p = 0.28). CONCLUSIONS: In Uro-EDIC women sexual activity rather than measures of diabetes control and complications was the main risk factor for urinary tract infection. The prevalence of cystitis was similar to that in nondiabetic women participants in NHANES III.

Detection of Mycoplasma genitalium in women with laparoscopically diagnosed acute salpingitis.

OBJECTIVES: Mycoplasma genitalium has been associated with cervicitis, endometritis, and tubal factor infertility. Because the ability of this bacterium to ascend and infect the fallopian tube remains undefined, we performed an investigation to determine the prevalence of M genitalium in fallopian tube, endometrial, and cervical specimens from women laparoscopically diagnosed with acute salpingitis in Nairobi, Kenya. METHODS: Women presenting with pelvic inflammatory disease were laparoscopically diagnosed with salpingitis. Infection with M genitalium in genital specimens was determined by polymerase chain reaction (PCR). RESULTS: Of 123 subjects with acute salpingitis, M genitalium was detected by PCR in the cervix and/or endometrium in nine (7%) participants, and in a single fallopian tube specimen. In addition, those infected with M genitalium were more often HIV infected than women not infected by M genitalium (seven of nine (78%) v 42 of 114 (37%), p<0.03). CONCLUSIONS: M genitalium is able to ascend into the fallopian tube, but its association with tubal pathology requires further investigation.

Susceptibility of Chlamydia trachomatis to excipients commonly used in topical microbicide formulations.

Commonly used "inactive" pharmaceutical excipients were tested in a previously developed minimum cidal concentration assay to assess their ability to kill Chlamydia trachomatis topically. Sixteen excipients were evaluated in these studies under various conditions. A range of activities was found among the excipients that could be tested in our assay system.

In vitro microbicidal activities of cecropin peptides D2A21 and D4E1 and gel formulations containing 0.1 to 2% D2A21 against Chlamydia trachomatis.

Topically applied microbicides that eradicate pathogens at the time of initial exposure represent a powerful strategy for the prevention of sexually transmitted infections. To aid in the further development of an effective topical microbicide, we assessed the minimum cidal concentration (MCC) of two cecropin peptides, D2A21 and D4E1, and gel formulations containing 0.1 to 2% D2A21 against Chlamydia trachomatis in vitro. The MCC of peptide D2A21 was 5 microM (18.32 microg/ml), and that of peptide D4E1 was 7.5 microM (21.69 microg/ml). The MCC of gel formulations containing 2% D2A21 was 0.2 mM (0.7 mg/ml), and that of gel formulations containing 0.5% D2A21 was 0.2 mM (0.7 mg/ml). There was no significant variation in the results when two different C. trachomatis strains were tested, and the addition of 10% human blood did not significantly alter the MCCs. pH values above and below 7 reduced the activity of the D2A21 peptide alone, but the MCC of the 2% D2A21 gel formulation was only slightly altered at the various pHs tested. Ultrastructural studies indicated that C. trachomatis membranes were disrupted after D2A21 exposure, resulting in leakage of the cytoplasmic contents. These in vitro results suggest that these cecropin peptides may be an effective topical microbicide against C. trachomatis and support the need for further evaluation.

Quantitative culture of Chlamydia trachomatis: relationship of inclusion-forming units produced in culture to clinical manifestations and acute inflammation in urogenital disease.

The relationship of Chlamydia trachomatis inclusion-forming units in quantitative culture to clinical manifestations and inflammation in urogenital disease was assessed in 1179 patients attending a sexually transmitted diseases clinic. In women, greater inclusion-forming unit counts were associated with cervical mucopus (3000 vs. 450 ifu), amount and character of cervical discharge, > or =30 polymorphonuclear cells (PMNL) per high-power field (hpf) on Gram stain (2050 vs. 320 ifu), and diagnoses of mucopurulent cervicitis (MPC; 2550 vs. 300 ifu) and pelvic inflammatory disease (PID; 3000 vs. 578 ifu). In men, greater inclusion-forming unit counts were associated with urethral discharge (85 vs. 44 ifu), amount and character of discharge, and > or =10 PMNL/hpf (95 vs. 50 ifu). These associations persisted on multivariate analysis. Thus, chlamydial replication is associated with MPC and PID in women, urethritis in men, and inflammation in both. Since infections with high inclusion counts may be the most transmissible, identification and treatment of patients with these chlamydia-associated syndromes is important in control programs.

Epidemiology and clinical manifestations of unique Chlamydia trachomatis isolates that occupy nonfusogenic inclusions.

Unique Chlamydia trachomatis strains characterized by multiple nonfusing inclusions were recently described. These strains lack evidence of the protein IncA in the inclusion membrane and have mutations in the incA gene. This study evaluated the epidemiology and clinical manifestations of patients infected with nonfusing mutant strains (case patients) and compared them with patients infected with wild-type fusing strains (control subjects). Both male and female case patients had fewer signs of infection than did control subjects (P=.016 and P=.019, respectively). Female case patients also had fewer symptoms of infection (P=.02). Median inclusion-forming unit (ifu) counts were lower in male and female case patients (P=.045 and P=.135, respectively). Thus, nonfusing strains of C. trachomatis more often produce subclinical infections than do normal fusing strains and have lower median ifu counts. From a prevention perspective, the data underscore the importance of screening programs to detect and treat inapparent C. trachomatis infection.

Evaluation of laboratory testing methods for Chlamydia trachomatis infection in the era of nucleic acid amplification.

Diagnostic tests presently available for Chlamydia trachomatis have widely varying performance characteristics. To assess evolving laboratory testing practices since the introduction of nucleic acid amplification tests (NAAT), we surveyed laboratories in Washington State about their testing practices in 1998 and compared our findings to a similar survey conducted in 1995. Laboratory directors of 61 (87%) of 70 laboratories performing chlamydial tests in 1998 returned a survey. Between 1995 and 1998, 36 laboratories discontinued chlamydial testing, and the total number of laboratories performing tests in the state decreased from 92 to 70, a 24% decline. Of the 36 laboratories that discontinued testing, 25 (69%) had previously used rapid tests. While no laboratory routinely used NAAT in 1995, ligase chain reaction (LCR) was used in 23% of laboratories in 1998 and accounted for 113,624 (36%) of the 318,133 tests performed that year. Among the remaining 204,509 tests performed in 1998, other tests employed included DNA probe (29%), enzyme immunoassay (20%), culture (12%), direct fluorescent antibody assays (3%), and rapid tests (<1%). The majority (65%) of tests performed in 1998 using technologies other than LCR or culture were done in laboratories that did more than 10,000 tests. Cost and loss of revenue to laboratories were the most frequently cited reasons for not adopting NAAT. We conclude that in Washington State, NAAT have been rapidly adopted in larger laboratories, but most patients are still tested with much less sensitive technologies. Financial constraints represent the major barrier to more widespread use of DNA amplification tests.

Patient-initiated treatment of uncomplicated recurrent urinary tract infections in young women.

BACKGROUND: Recurrent urinary tract infections (UTIs) are a common outpatient problem, resulting in frequent office visits and often requiring the use of prophylactic antimicrobial agents. Patient-initiated treatment of recurrent UTIs may decrease antimicrobial use and improve patient convenience. OBJECTIVE: To determine the safety and feasibility of patient-initiated treatment of recurrent UTIs. DESIGN: Uncontrolled, prospective clinical trial. SETTING: University-based primary health care clinic. PARTICIPANTS: Women at least 18 years of age with a history of recurrent UTIs and no recent pregnancy, hypertension, diabetes, or renal disease. INTERVENTION: After self-diagnosing UTI on the basis of symptoms, participating women initiated therapy with ofloxacin or levofloxacin. MEASUREMENTS: Accuracy of self-diagnosis determined by evidence of a definite (culture-positive) or probable (sterile pyuria and no alternative diagnosis) UTI on pretherapy urinalysis and culture. Women with a self-diagnosis of UTI that was not microbiologically confirmed were evaluated for alternative diagnoses. Post-therapy interviews and urine cultures were used to assess clinical and microbiological cure rates, adverse events, and patient satisfaction. RESULTS: 88 of 172 women self-diagnosed a total of 172 UTIs. Laboratory evaluation showed a uropathogen in 144 cases (84%), sterile pyuria in 19 cases (11%), and no pyuria or bacteriuria in 9 cases (5%). Clinical and microbiological cures occurred in 92% and 96%, respectively, of culture-confirmed episodes. No serious adverse events occurred. CONCLUSION: Adherent women can accurately self-diagnose and self-treat recurrent UTIs.

Increasing antimicrobial resistance and the management of uncomplicated community-acquired urinary tract infections.

Community-acquired urinary tract infections (UTIs) are among the most common bacterial infections in women. Therapy for these infections is usually begun before results of microbiological tests are known. Furthermore, in women with acute uncomplicated cystitis, empirical therapy without a pretherapy urine culture is often used. The rationale for this approach is based on the highly predictable spectrum of etiologic agents causing UTI and their antimicrobial resistance patterns. However, antimicrobial resistance among uropathogens causing community-acquired UTIs, both cystitis and pyelonephritis, is increasing. Most important has been the increasing resistance to trimethoprim-sulfamethoxazole (TMP-SMX), the current drug of choice for treatment of acute uncomplicated cystitis in women. What implications do these trends have for treatment of community-acquired UTIs? Preliminary data suggest that clinical cure rates may be lower among women with uncomplicated cystitis treated with TMP-SMX when the infecting pathogen is resistant to TMP-SMX. Women with pyelonephritis also have less bacterial eradication and lower clinical cure rates when treated with TMP-SMX for an infection that is resistant to the drug. Therefore, in the outpatient setting, identifying risk factors for TMP-SMX resistance and knowing the prevalence of TMP-SMX resistance in the local community are important steps in choosing an appropriate therapeutic agent. When choosing a treatment regimen, physicians should consider such factors as in vitro susceptibility, adverse effects, cost-effectiveness, and selection of resistant strains. Using a management strategy that takes these variables into account is essential for maintaining the safety and efficacy of treatment for acute UTI.

Antimicrobial resistance among uropathogens that cause community-acquired urinary tract infections in women: a nationwide analysis.

Current recommendations for empirical therapy for community-acquired urinary tract infection (UTI) in women hinge on knowledge of antimicrobial susceptibility patterns in the geographic region of the practitioner. We conducted a survey of antimicrobial susceptibilities of 103,223 isolates recovered from urine samples that were obtained in 1998 from female outpatients nationally and within 9 geographic regions in the United States. Resistance of Escherichia coli isolates to trimethoprim-sulfamethoxazole varied significantly according to geographic region, ranging from a high of 22% in the western United States to a low of 10% in the Northeast (P<.001). There were no clinically significant age-related differences in the susceptibility of E. coli to any of the study drugs, but the susceptibility to fluoroquinolones of non-E. coli isolates that were recovered from women who were aged >50 years was significantly lower than that of isolates recovered from younger women (P<.001). The in vitro susceptibility of uropathogens in female outpatients varies according to age and geographic region.

Urine-based screening for Chlamydia trachomatis in men attending sexually transmitted disease clinics.

BACKGROUND: Nucleic acid-amplified tests for Chlamydia trachomatis are accurate but costly. Screening strategies for asymptomatic men are needed. GOAL: To assess C trachomatis screening strategies for asymptomatic males. STUDY DESIGN: Men attending a sexually transmitted disease clinic were tested for C trachomatis with ligase chain reaction and culture, and for urethral inflammation with urine leukocyte esterase and urethral Gram stain. RESULTS: C trachomatis prevalence was 5.5% among 1,625 asymptomatic men. Ligase chain reaction increased detection by 49% among men without urethral inflammation. An age of younger than 25 years and urethral inflammation were associated with positive ligase chain reaction results. The negative predictive value of urine leukocyte esterase was highest among older men, but urethral Gram stain was equally sensitive in predicting infection regardless of age. An age of younger than 30 years or urethral inflammation identified the highest proportion of infections (92%) and reduced the percentage of men screened by 43%. CONCLUSIONS: Urine ligase chain reaction increased C trachomatis detection, particularly among men without urethral inflammation. Testing all asymptomatic men younger than 30 years is optimal, whereas negative urine leukocyte esterase or urethral Gram stain results in men 30 years or older support no testing.

Adequacy of fellowship training: results of a survey of recently graduated fellows.

The adequacy of fellowship training in the field of infectious diseases was assessed by means of a survey of recently graduated fellows. Surveys were mailed to all individuals who had passed the American Board of Internal Medicine's board certification examination in infectious diseases since 1992. A total of 666 completed surveys were returned by the deadline (response rate, 36%). Although most recent graduates thought that training in the standard components of clinical infectious diseases was adequate, only 50% thought that training in infection control was adequate. Fewer than 1 in 3 believed that they had received adequate training in the business aspects of infectious diseases practice. The adequacy and duration of research training were linked to ultimate career choice. These results form the basis for the Infectious Diseases Society of America's new initiatives to assist with more-diversified and relevant fellowship training.

Pneumonia and sepsis due to fluoroquinolone-resistant Capnocytophaga gingivalis after autologous stem cell transplantation.

Human oral Capnocytophaga species have been only rarely described as a cause of sepsis in patients following stem cell or marrow transplantation, and pneumonia has not been reported in this setting. In addition, fluoroquinolone resistance is rarely seen in these species, and has never been reported in C. gingivalis. We report a case of pneumonia (confirmed by culture of bronchoalveolar lavage fluid) and sepsis due to fluoroquinolone- resistant Capnocytophaga gingivalis in a patient following autologous stem cell transplantation, who responded to treatment with linezolid and metronidazole. Capnocytophaga infections should be considered in patients with fever following stem cell or marrow transplantation, especially those with neutropenia and mucositis. Susceptibility testing is needed given the existence of multidrug-resistant isolates.