Molecular epidemiology of Ornithobacterium rhinotracheale

Ornithobacterium rhinotracheale is a recently described gram-negative rod-shaped bacterium associated with respiratory tract infections in poultry. In order to determine the molecular epidemiology of this bacterium, we characterized 55 O. rhinotracheale isolates from eight countries on four continents by multilocus enzyme electrophoresis (MLEE), repetitive sequence based-PCR (rep-PCR), and 16S rRNA gene sequencing. MLEE discriminated the O. rhinotracheale isolates into six electrophoretic types (ETs), of which only three ETs were recovered from domesticated poultry. The 16S rRNA gene sequence and rep-PCR analyses confirmed the results obtained by MLEE and indicated limited heterogeneity among isolates of O. rhinotracheale recovered from poultry. Taken together, the results of our analysis demonstrate that the majority of O. rhinotracheale isolates recovered from domesticated poultry throughout the world are represented by a small group of closely related clones and suggest that the bacterium was recently introduced to domesticated poultry from wild bird populations.     

Molecular epidemiology of hepatitis C in Australia

The aim of this study was to determine the distribution of hepatitis C virus (HCV) genotypes in Australian patients with hepatitis C and to identify factors associated with particular genotypes. Serum isolates of HCV-RNA were genotyped using a commercial oligonucleotide hybridization (line probe) assay. Relationships between demographic factors, mode of HCV transmission and HCV genotype were assessed by logistic regression analysis. Among 463 patients with hepatitis C, 425 tested positive for HCV-RNA and a single HCV genotype was identified in 420 cases. The patients' places of birth were Australia or New Zealand (62%), Asia (13%), Europe (12%), Mediterranean (6%), Middle East (6%) and other countries (< 1%). The most common genotypes were type 1 (52%) or type 3 (32%); type 2 (9.3%), type 4 (5.5%) and type 6 (1.7%) were less common. Patients with genotype 1b were older (48 +/- 13 years, P< 0.001) and patients with genotype 3 were younger than the remaining patients (37 +/- 11 years vs 42 +/- 12 years, P< 0.001). Among type 1 isolates, 1b was more common for patients born outside Australia compared with those born in Australia (50% vs 13%, P< 0.001) whereas non-1b subtypes were more common among Australian-born patients. Likewise, 21 of 23 (91%) patients with type 4 were from Egypt and six of seven (86%) with type 6 were from Vietnam. The relative importance of parenteral risk factors for HCV also varied according to geographic origin. Thus, a definite risk factor for HCV acquisition was identified in > 95% of Australian-born patients, but in only 33% of Asian or Mediterranean-born patients. Logistic regression analysis indicated that region of birth and risk factor (intravenous drug use or not) would allow 98% of type 4 cases and 76% of type 1b cases to be identified correctly. In summary, region of birth, patterns of migration over time and risk factors for transmission of HCV interact to determine the distribution of HCV genotypes in a multi-racial community like Australia.     

Molecular epidemiology of tuberculosis in Denmark in 1992

The incidence of tuberculosis (TB) is increasing all over the world, including in countries with a high standard of living and good social security. Denmark represents such a region. Furthermore, it is a small country (5 million inhabitants) with a long tradition in TB control, including a centralization of the bacteriological diagnostic facility. The present study was intended to analyze the transmission of Mycobacterium tuberculosis in a country in which TB has low endemicity by a combination of conventional epidemiological approaches and DNA fingerprinting techniques, whereby individual bacterial strains can be traced. M. tuberculosis isolates from 92% of all new cases of bacteriologically verified TB in Denmark during 1992 were subjected to IS6110 DNA fingerprinting to visualize the DNA restriction fragment length polymorphism (RFLP) patterns of the isolated strains. The data obtained from the RFLP analyses were interpreted by using demographic data, such as age, sex, ethnicity, and residence, for the patients. The risk factors among the patients for being part of an active chain of transmission, as opposed to demonstrating reactivation of a previously acquired latent infection, were estimated by statistical analyses. The magnitude of TB transmission in 1992 in Denmark was determined, and transmitted infections were shown to comprise at least one quarter of the total number of cases. Almost half of the TB cases involved patients of foreign origin. However, most of these isolates showed unique DNA fingerprint patterns and were rarely part of an active chain of transmission. The major chains of recent transmission were localized to distinct geographical regions in the country. TB is frequent among immigrants, especially from Asia and Africa, but it is apparently readily suspected, diagnosed, and treated by the health care system. Danish patients with pulmonary symptoms are not primarily suspected to have TB and, therefore, play an important role in recent TB transmission in Denmark.     

Molecular epidemiology and the new tuberculosis

DNA fingerprinting techniques now exist which identify specific strains of Mycobacterium tuberculosis. These techniques may be integrated with conventional epidemiologic approaches to better understand tuberculosis in its modern form. This paper reviews the lessons learned from this approach about the pathogenesis and epidemiology of tuberculosis. In addition, it speculates about the potential future applications of molecular epidemiology, including its use as an adjunct to conventional public health measures.     

Molecular identification and epidemiology of animal mycoplasmas

Molecular genetic techniques show a high potential for rapid and accurate identification of Mycoplasma species isolated from animals. An important field of application for such methods is the differentiation of species and/or subspecies which are phenotypically closely related, but which show significant differences in epidemiological impact. This need is particularly important for the mycoplasmas of the "mycoides group", which are phenotypically and phylogenetically very closely related. Molecular typing techniques based on 16S rRNA genes give straightforward phylogenetic answers on the species level. For more refined methods of subtyping at the subspecies level, the use of defined genes characteristic of certain Mycoplasma species or clusters is recommended. Genetic fingerprinting, especially insertion sequence typing has proved to be a valuable tool for subtyping and strain identification in particular of vaccine strains and for epidemiological investigations.     

Molecular epidemiology of Staphylococcus aureus and Enterococcus faecalis in endophthalmitis

Genomic DNA fingerprint analysis was performed on 39 Staphylococcus aureus and 28 Enterococcus faecalis endophthalmitis isolates collected from multiple clinical centers. Among 21 S. aureus genomic DNA fingerprint patterns identified, five clonotypes were recovered from multiple unrelated patients and accounted for 58.9% (23 of 39) of the isolates analyzed. Compared with strains having unique genomic DNA fingerprint patterns, the S. aureus clonotypes occurring more than once were more likely to result in visual acuities of 20/200 or worse (P = 0.036 [chi2 test]). In contrast to the S. aureus isolates, the E. faecalis endophthalmitis isolates were a clonally diverse population, enriched for the expression of a known toxin, cytolysin, which is plasmid encoded.     

Molecular epidemiology of bartonella infections in patients with bacillary angiomatosis-peliosis

BACKGROUND: Bacillary angiomatosis and bacillary peliosis are vascular proliferative manifestations of infection with species of the genus bartonella that occur predominantly in patients infected with the human immunodeficiency virus. Two species, B. henselae and B. quintana, have been associated with bacillary angiomatosis, but culture and speciation are difficult, and there has been little systematic evaluation of the species-specific disease characteristics. We studied 49 patients seen over eight years who were infected with bartonella species identified by molecular techniques and who had clinical lesions consistent with bacillary angiomatosis-peliosis. METHODS: In this case-control study, a standardized questionnaire about exposures was administered to patients with bacillary angiomatosis-peliosis and to 96 matched controls. The infecting bartonella species were determined by molecular techniques. RESULTS: Of the 49 patients with bacillary angiomatosis-peliosis, 26 (53 percent) were infected with B. henselae and 23 (47 percent) with B. quintana. Subcutaneous and lytic bone lesions were strongly associated with B. quintana, whereas peliosis hepatis was associated exclusively with B. henselae. Patients with B. henselae infection were identified throughout the study period and were epidemiologically linked to cat and flea exposure (P< or =0.004), whereas those with B. quintana were clustered and were characterized by low income (P=0.003), homelessness (P = 0.004), and exposure to lice (P= 0.03). Prior treatment with macrolide antibiotics appeared to be protective against infection with either species. CONCLUSIONS: B. henselae and B. quintana, the organisms that cause bacillary angiomatosis-peliosis, are associated with different epidemiologic risk factors and with predilections for involvement of different organs.     

Molecular characterization of a novel Rickettsia species from Ixodes scapularis in Texas

Triclosan (Irgasan) is a broad spectrum antimicrobial agent used in handsoaps, toothpastes, fabrics, and plastics. It inhibits lipid biosynthesis in Escherichia coli, probably by action upon enoyl reductase (FabI) (McMurry, L.M., Oethinger, M. and Levy, S.B. (1988) Nature 394, 531-532). We report here that overexpression of the multidrug efflux pump locus acrAB, or of marA or soxS, both encoding positive regulators of acrAB, decreased susceptibility to triclosan 2-fold. Deletion of the acrAB locus increased the susceptibility to triclosan approximately 10-fold. Four of five clinical E. coli strains which overexpressed marA or soxS also showed enhanced triclosan resistance. The acrAB locus was involved in the effects of triclosan upon both cell growth rate and cell lysis.     

Molecular epidemiology of oral treponemes associated with periodontal disease

Periodontitis, a disease responsible for tooth loss worldwide, is characterized by chronic inflammation of the periodontium, eventually leading to destruction of periodontal ligaments and supporting alveolar bone. Spirochetes, identified by dark-field microscopy as being the most predominant bacteria in advanced lesions, are thought to play a causative role. Various spirochetal morphotypes were observed, but most of these morphotypes are as yet uncultivable. To assess the role of these organisms we designed oligonucleotide probes for the identification of both cultivable and so far uncultivable spirochetes in periodontitis patients. Subgingival plaque specimens taken from diseased sites (n = 200) and healthy control sites (n = 44) from 53 patients with rapidly progressive periodontitis (RPP) were submitted to direct in situ hybridization or dot blot hybridization after prior amplification with eubacterial primers. Spirochetes were found in all patients, but their distributions varied considerably. Parallel use of oligonucleotide probes specific for cultivable or so far uncultivable treponemes suggested the presence of novel yet unknown organisms at a high frequency. These uncultivable treponemes were visualized by fluorescence in situ hybridization, and their morphologies, sizes, and numbers could be estimated. All RPP patients included in this study harbored oral treponemes that represent either novel species, e.g., Treponema maltophilum, or uncultivable phylotypes. Therefore, it is necessary to include these organisms in etiologic considerations and to strengthen efforts to cultivate these as yet uncultivable treponemes.     

Molecular epidemiology of group A streptococcus M type 1 infections

OBJECTIVE: To determine if the motility stimulants, caffeine (7 mM), pentoxifylline (3 mM), 2-deoxyadenosine (3 mM), and a combination of the three would induce hyperactivated (HA) motility. DESIGN: Controlled laboratory study of stimulants using cryopreserved semen from 10 donors at three time points. SETTING: The study was performed in the Andrology Laboratory at Prince Henry's Institute of Medical Research Clayton, Victoria, Australia. INTERVENTIONS: Stimulants in doses maximally effective for stimulation of motility were incubated with suspensions of previously cryopreserved sperm. MAIN OUTCOME MEASURES: Motility characteristics (curvilinear velocity [VCL], linearity [LIN], and maximum amplitude of lateral head displacement [ALHmax]) were derived using the single cell track facility of the CellSoft computer-automated semen analyzer (Cryo Resources Ltd., Montgomery, NY). Videotapes were visually inspected, and 125 sperm cell trajectories exhibiting characteristic HA behavior were identified. The HA motility thresholds (5th or 95th centiles) were as follows: VCL > 74 microns/s, LIN < 74%, and ALHmax > 4.7 microns. Cells with motility characteristics outside these limits were regarded as HA. The significance of the effect of the stimulants on the proportion of sperm exhibiting HA was examined by Poisson regression analysis. RESULTS: Sperm washing (removal of the cells from the cryoprotectant) and swim-up caused significant changes in the VCL and straight line velocity and a twofold (5.6% to 11.9%) increase in the proportion of HA cells. In the presence of motility stimulants, the proportion of HA cells was significantly increased threefold (11.9% to 32.5%) above that seen in the control washed sperm. Hyperactivated motility declined after 1 to 2 hours, but 2-deoxyadenosine demonstrated a prolonged effect. CONCLUSION: These motility stimulants that affect adenosine 3':5' monophosphate in human sperm stimulate cyclic hyperactivation.     

Molecular epidemiology of recent belgian isolates of Neisseria meningitidis serogroup B

In Belgium an increase in the incidence of meningococcal disease has been noted since the early 1990s. Four hundred twenty clinical strains isolated during the period from 1990 to 1995, along with a set of 30 European reference strains, and 20 Dutch isolates were examined by random-primer and repetitive-motif-based PCR. A subset was investigated by multilocus enzyme electrophoresis and pulsed-field gel electrophoresis. The data were compared with results obtained by serotyping (M. Van Looveren, F. Carion, P. Vandamme, and H. Goossens, Clin. Microbiol. Infect. 4:224-228, 1998). Both phenotypic and molecular epidemiological data suggest that the lineage III of Neisseria meningitidis, first encountered in The Netherlands in about 1980, has been introduced in Belgium. The epidemic clone, as defined by oligonucleotide D8635-primed PCR, encompasses mainly phenotypes B:4:P1.4 and B:nontypeable:P1.4, but strains with several other phenotypes were also encountered. Therefore, serotyping alone would underestimate the prevalence of the epidemic clone.     

Molecular epidemiology of sporadic (endemic) serogroup C meningococcal disease

Understanding the basis of sporadic (endemic) meningococcal disease may be critical to prevention of meningococcal epidemic outbreaks and to understanding fluctuations in incidence. Active, prospective, population-based surveillance and molecular epidemiologic techniques were used to study sporadic serogroup C meningococcal disease in a population of 2.34 million persons (Atlanta area). During 1988-1994, in which no outbreaks or case clusters were reported, 71 patients developed sporadic serogroup C meningococcal disease (annual incidence, 0.51/100,000). Eighty-three percent of patients were >2 years old. By multilocus enzyme electrophoresis, pulsed-field gel electrophoresis, and serotyping, 84% (52/62) of the isolates available for study were identical or closely related members of the electrophoretic type 37 (ET 37) complex responsible for multiple serogroup C outbreaks in the United States in the 1990s. Sporadic disease caused by 9 clonal strains occurred over periods up to 4 years and accounted for 45% (28/62) of cases. Sporadic serogroup C meningococcal disease was most often due to a limited number of related strains that appear to slowly circulate in the population.     

Molecular epidemiology of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae isolates in a district hospital in Taiwan

Thirty-one of 104 clinical isolates of Klebsiella pneumoniae collected over a period of 8 months were found to be putative extended-spectrum beta-lactamase (ESBL) producers. Isoelectric focusing and an iodine overlay agar method were used for preliminary identification of the ESBLs. They were further identified by DNA sequencing. Seventy-one percent of the isolates were found to produce SHV-5. The variation in the ESBL patterns of these isolates was slight, with only five patterns being identified. The strains were typed by pulsed-field gel electrophoresis (PFGE), and 16 different genotypes were identified. When the PFGE patterns were analyzed by the algorithmic clustering method called the unweighted-pair group method using arithmetic averages, five clusters were found. However, significant genetic variations were found among 11 isolates and between each cluster. A plasmid of 36 kb was found in all clinical isolates and in the transconjugants. Our results indicate that the increase in the number of ESBL-producing K. pneumoniae isolates in this hospital is due mainly to the dissemination of a resistance plasmid rather than to the clonal spread of a few epidemic strains.     

Molecular genetics and the epidemiology of common mental disorders: new opportunities

We present results of a study on isolation and endogamy in San Antonio de Los Altos a Venezuelan town founded in 1683 by 41 Spanish families of Canarian origin. The study was done with the civil marital records registered during the years 1905 to 1981. The results show that the same few surnames cover an important proportion of individuals in all the periods studied, which suggests that these belong to the traditional families. An important endogamy is revealed through the frequencies of surnames, birth places of spouses, and the isonymy analysis, which decrease after 1960 with the rupture of the isolate. The higher Fn values and the predominance of some types of isonymy suggests the existence of preferential unions. Geographic isolation and some economic factors, such as the property of the land, could have favoured the isonymy and endogamy observed among surnames of the founders of the town.