Carl F. Marrs

Identification of Risk Factors for Extrapulmonary Tuberculosis

The proportion of extrapulmonary tuberculosis cases in the United States has increased from 16% of tuberculosis cases, in 1991, to 20%, in 2001. To determine associations between the demographic, clinical, and life style characteristics of patients with tuberculosis and the occurrence of extrapulmonary tuberculosis, a retrospective case-control study was conducted. This study included 705 patients with tuberculosis, representing 98% of the culture-proven cases of tuberculosis in Arkansas from 1 January 1996 through 31 December 2000. A comparison between 85 patients with extrapulmonary tuberculosis (case patients) and 620 patients with pulmonary tuberculosis (control patients) showed women (OR, 1.98; 95% CI, 1.25-3.13), non-Hispanic blacks (OR, 2.38; 95% CI, 1.42-3.97), and HIV-positive persons (OR, 4.93; 95% CI, 1.95-12.46) to have a significantly higher risk for extrapulmonary tuberculosis than men, non-Hispanic whites, and HIV-negative persons. This study expands the knowledge base regarding the epidemiology of extrapulmonary tuberculosis and enhances our understanding of the relative contribution of host-related factors to the pathogenesis of tuberculosis.

Prevalence of Antibiotic Resistance in Drinking Water Treatment and Distribution Systems

ABSTRACT The occurrence and spread of antibiotic-resistant bacteria (ARB) are pressing public health problems worldwide, and aquatic ecosystems are a recognized reservoir for ARB. We used culture-dependent methods and quantitative molecular techniques to detect and quantify ARB and antibiotic resistance genes (ARGs) in source waters, drinking water treatment plants, and tap water from several cities in Michigan and Ohio. We found ARGs and heterotrophic ARB in all finished water and tap water tested, although the amounts were small. The quantities of most ARGs were greater in tap water than in finished water and source water. In general, the levels of bacteria were higher in source water than in tap water, and the levels of ARB were higher in tap water than in finished water, indicating that there was regrowth of bacteria in drinking water distribution systems. Elevated resistance to some antibiotics was observed during water treatment and in tap water. Water treatment might increase the antibiotic resistance of surviving bacteria, and water distribution systems may serve as an important reservoir for the spread of antibiotic resistance to opportunistic pathogens.

Wastewater treatment contributes to selective increase of antibiotic resistance among Acinetobacter spp.

The occurrence and spread of multi-drug resistant bacteria is a pressing public health problem. The emergence of bacterial resistance to antibiotics is common in areas where antibiotics are heavily used, and antibiotic-resistant bacteria also increasingly occur in aquatic environments. The purpose of the present study was to evaluate the impact of the wastewater treatment process on the prevalence of antibiotic resistance in Acinetobacter spp. in the wastewater and its receiving water. During two different events (high-temperature, high-flow, 31 degrees C; and low-temperature, low-flow, 8 degrees C), 366 strains of Acinetobacter spp. were isolated from five different sites, three in a wastewater treatment plant (raw influent, second effluent, and final effluent) and two in the receiving body (upstream and downstream of the treated wastewater discharge point). The antibiotic susceptibility phenotypes were determined by the disc-diffusion method for 8 antibiotics, amoxicillin/clavulanic acid (AMC), chloramphenicol (CHL), ciprofloxacin (CIP), colistin (CL), gentamicin (GM), rifampin (RA), sulfisoxazole (SU), and trimethoprim (TMP). The prevalence of antibiotic resistance in Acinetobacter isolates to AMC, CHL, RA, and multi-drug (three antibiotics or more) significantly increased (p<0.01) from the raw influent samples (AMC, 8.7%; CHL, 25.2%; RA, 63.1%; multi-drug, 33.0%) to the final effluent samples (AMC, 37.9%; CHL, 69.0%; RA, 84.5%; multi-drug, 72.4%), and was significantly higher (p<0.05) in the downstream samples (AMC, 25.8%; CHL, 48.4%; RA, 85.5%; multi-drug, 56.5%) than in the upstream samples (AMC, 9.5%; CHL, 27.0%; RA, 65.1%; multi-drug, 28.6%). These results suggest that wastewater treatment process contributes to the selective increase of antibiotic resistant bacteria and the occurrence of multi-drug resistant bacteria in aquatic environments.

Both urinary and rectal Escherichia coli isolates are dominated by strains of phylogenetic group B2.

ABSTRACT To compare the genetic structures of uropathogenic and commensal Escherichia coli populations, a total of 181 urinary and rectal E. coli isolates were classified into intraspecies phylogenetic groups by PCR amplifications of phylogenetic markers. The genetic variability of these isolates within phylogenetic groups was further assessed by enterobacterial repetitive intergenic consensus (ERIC) typing. The distributions of 10 known virulence factors were also examined. In contrast with most reports, phylogenetic group B2 not only accounted for the majority of urinary isolates from young women with urinary tract infections (69%) but also was the dominant group among the rectal isolates from healthy young women (48%). Such difference may be explained by geographic variation, difference in host population characteristics, or differences in sampling method, or a combination of the three. Strains with known virulence factors most frequently belonged to phylogenetic groups B2 and D. Additionally, group B2 and D rectal isolates were more heterogeneous than urinary isolates. Two subclusters existed within group B2 strains by ERIC typing. These subclusters were not evenly distributed between rectal and urine isolates and differed in virulence gene distribution.

Association between Mycobacterium tuberculosis Beijing/W Lineage Strain Infection and Extrathoracic Tuberculosis: Insights from Epidemiologic and Clinical Characterization of the Three Principal Genetic Groups of M. tuberculosis Clinical Isolates

ABSTRACT Clinical strains of Mycobacterium tuberculosis can be divided into three principal genetic groups based on the single-nucleotide polymorphisms at the katG gene codon 463 and the gyrA gene codon 95. One subgroup of genetic group 1, the Beijing/W lineage, has been widely studied because of its worldwide distribution and association with outbreaks. In order to increase our understanding of the clinical and epidemiological relevance of the genetic grouping of M. tuberculosis clinical strains and the Beijing/W lineage, we investigated the genetic grouping of 679 clinical isolates of M. tuberculosis, representing 96.3% of culture-confirmed tuberculosis cases diagnosed in Arkansas between January 1996 and December 2000 using PCR and DNA sequencing. We assessed the associations of infections by different genetic groups of M. tuberculosis strains and infection by the Beijing/W lineage strains with the clinical and epidemiological characteristics of the patients using chi-square tests and multivariate logistic regression analysis. Of the 679 study isolates, 676 fell into one of the three principal genetic groups, with 63 (9.3%) in group 1, 438 (64.8%) in group 2, and 175 (25.9%) in group 3. After adjusting for potential confounding of age, gender, race/ethnicity, human immunodeficiency virus serostatus, and plcD genotype in a multivariate logistic regression model, patients infected by the Beijing/W lineage isolates were nearly three times as likely as patients infected with the non-Beijing/W lineage isolates to have an extrathoracic involvement (odds ratio [95% confidence interval], 2.85 [1.33, 6.12]). Thus, the Beijing/W lineage strains may have some special biological features that facilitate the development of extrathoracic tuberculosis.

Relationships of Nontypeable Haemophilus influenzae Strains to Hemolytic and Nonhemolytic Haemophilus haemolyticus Strains

ABSTRACT Haemophilus influenzae is both a human respiratory pathogen and pharyngeal commensal, while H. haemolyticus, the closest phylogenetic relative of H. influenzae, is arguably a strict pharyngeal commensal. A hemolytic phenotype has historically differentiated H. haemolyticus from H. influenzae, but the recent recognition of significant nonhemolytic H. haemolyticus colonization has decreased this traits resolvability. Given this and the potential of recombination between the species, we examined the distribution of microbiologic and molecular traits between collections of H. influenzae and H. haemolyticus strains separated within a dendrogram obtained by multilocus sequence analysis (MLSA). All strains hybridizing with a probe to iga, a gene encoding an immunoglobulin A protease of H. influenzae, clustered apart from strains that did not hybridize with the probe. Other traits also segregated significantly along this division, suggesting a separation of the species. Of note, the LOS genes licA, lic2A, and lgtC of H. influenzae were approximately 2, 6, and 54 times, respectively, more prevalent in H. influenzae than in H. haemolyticus. In contrast to species separation, interspecies recombination was evidenced by the inability of single gene sequences to phylogenetically separate the species and by the “fuzzy” distribution of some species-specific traits across the species dividing line. Together, these data support the historically accurate and pragmatic division of these species while recognizing their potential for recombination. Future comparative genomic studies identifying common and distinctive genes could be useful in evaluating their role in the commensal or virulent growth, respectively, of H. influenzae.

Diversity and sharing of Haemophilus influenzae strains colonizing healthy children attending day-care centers

Background. Children attending day-care centers (DCCs) are at risk for Haemophilus influenzae nasopharyngeal colonization and acute otitis media. The degree to which a given strain circulates within a day-care center and the heterogeneity of strains among DCCs in a geographic area are not well-characterized. This study describes the prevalence rates of H. influenzae colonization in a large number of children attending day-care centers and examines the genetic diversity of colonizing strains and the degree of sharing among children. Methods. Throat cultures were collected from 198 healthy children <3 years old attending 16 day-care centers in Michigan. All H. influenzae isolates were genetically typed by enterobacterial repetitive intergenic consensus PCR as the initial screening technique to identify unique strains within each child. Pulsed field gel electrophoresis was used subsequently to examine the genetic diversity of strains between children. Results. There were 127 (64%) children colonized with H. influenzae. Wide variation in rates of colonization was identified among day-care centers (0 to 95%). A total of 179 genetically unique H. influenzae strains were isolated, and 47 children (37%) were colonized with 2 or more genetically distinct H. influenzae organisms. Evidence of sharing of the same strain in different children was found in 13 of 15 colonized DCCs and 23% of genotypically unique strains were shared. Conclusion. The degree of sharing of H. influenzae among children in this study suggests transmission of these potentially pathogenic microorganisms in day-care centers.

Molecular Epidemiology of 3 Putative Virulence Genes for Escherichia coli Urinary Tract Infection—usp, iha, and iroNE. coli

This study describes the epidemiological association of 3 putative genes for virulence of uropathogenic Escherichia coli; uropathogenic specific protein (usp), a Vibrio cholerae zot gene homologue; IrgA homologue adhesin (iha), a nonhemagglutinating adhesin; and iroN(E. coli), a catechole siderophore receptor homologue. We compared the relative frequency in urinary tract infection (UTI) isolates (n=508), compared with non-UTI isolates (n=416). iroN(E. coli) occurred 2.1-3.6 times more frequently in UTI isolates than in rectal isolates (P=1.1x10-18 to P=2.7x10-5) and was associated with several uropathogenic virulence genes found on pathogenicity islands. usp occurred more frequently in isolates from patients with pyelonephritis (P=3.6x10-9), in periurethral isolates (P=.001), and in isolates from patients with UTI who were aged 40-65 years (P=.004), when compared with the rectal isolates; iha was not associated with UTI in this study.

Choosing an appropriate bacterial typing technique for epidemiologic studies

A wide variety of bacterial typing systems are currently in use that vary greatly with respect to the effort required, cost, reliability and ability to discriminate between bacterial strains. No one technique is optimal for all forms of investigation. We discuss the desired level of discrimination and need for a biologic basis for grouping strains of apparently different types when using bacterial typing techniques for different epidemiologic applications: 1) confirming epidemiologic linkage in outbreak investigations, 2) generating hypotheses about epidemiologic relationships between bacterial strains in the absence of epidemiologic information, and 3) describing the distributions of bacterial types and identifying determinants of those distributions. Inferences made from molecular epidemiologic studies of bacteria depend upon both the typing technique selected and the study design used; thus, choice of typing technique is pivotal for increasing our understanding of the pathogenesis and transmission, and eventual disease prevention.

Pharyngeal Colonization Dynamics of Haemophilus influenzae and Haemophilus haemolyticus in Healthy Adult Carriers

ABSTRACT Haemophilus influenzae is an important cause of respiratory infections, including acute otitis media, sinusitis, and chronic bronchitis, which are preceded by asymptomatic H. influenzae colonization of the human pharynx. The aim of this study was to describe the dynamics of pharyngeal colonization by H. influenzae and an intimately related species, Haemophilus haemolyticus, in healthy adults. Throat specimens from four healthy adult carriers were screened for Haemophilus species; 860 isolates were identified as H. influenzae or H. haemolyticus based on the porphyrin test and on dependence on hemin and NAD for growth. Based on tests for hemolysis, for the presence of the 7F3 epitope of the P6 protein, and for the presence of iga in 412 of the isolates, 346 (84%) were H. influenzae, 47 (11%) were H. haemolyticus, 18 (4%) were nonhemolytic H. haemolyticus, and 1 was a variant strain. Carriers A and B were predominantly colonized with nontypeable H. influenzae, carrier C predominantly with b−H. influenzae mutants, and carrier D with H. haemolyticus. A total of 358 H. influenzae and H. haemolyticus isolates were genotyped by pulsed-field gel electrophoresis (PFGE) following SmaI or EagI digestion of their DNA, and the carriers displayed the following: carrier A had 11 unique PFGE genotypes, carrier B had 15, carrier C had 7, and carrier D had 10. Thus, adult H. influenzae and H. haemolyticus carriers are colonized with multiple unique genotypes, the colonizing strains exhibit genetic diversity, and we observed day-to-day and week-to-week variability of the genotypes. These results appear to reflect both evolutionary processes that occur among H. influenzae isolates during asymptomatic pharyngeal carriage and sample-to-sample collection bias from a large, variable population of colonizing bacteria.