Non-pathogenic Actinobacillus isolates antigenically and biochemically similar to Actinobacillus pleuropneumoniae: a novel species?

Two unusual Actinobacillus isolates were recovered from pigs with no clinical signs, no lesions and no history of swine pleuropneumonia. Two representative strains (9953L55 and 0347) analyzed in this study were initially biochemically and antigenically identified as A. pleuropneumoniae serotypes 1 and 9, respectively, by traditional identification methods. Both strains presented, however, negative results with three A. pleuropneumoniae-specific PCR tests and revealed in particular the absence of the apxIV toxin genes. However, both strains produced and secreted ApxII toxin although they only harbored the toxin genes apxIICA, which is an uncommon feature for any of the known A. pleuropneumoniae serotypes. Upon experimental inoculation of pigs, these strains proved to be totally non-pathogenic. Animals infected with one of the strains produced antibodies that cross-react with A. pleuropneumoniae serotypes 1-9-11-specific LC-LPS ELISA. Phylogenetic analysis based on 16S rRNA gene sequence analysis revealed that these strains form a separate phylogenetic group that is distinct from other Actinobacillus species and is particularly different from A. pleuropneumoniae.

Characterization of Aqx and its operon: the hemolytic RTX determinant of Actinobacillus equuli

Actinobacillus equuli, a member of the family Pasteurellaceae is the etiologic agent of a frequently lethal septicemia in neonatal foals as well as other more chronic diseases like arthritis, pleuritis, pneumonia or peritonitis. It may also be isolated from the oral cavity of healthy horses. Hemolytic isolates of A. equuli are known but so far no virulence determinants have been described for this bacterial species. By screening hemolytic A. equuli strains with specific gene probes, a hemolysin, designated Aqx (A. equuli RTX (repeats in the structural toxin)) was identified. This hemolysin was shown to be an RTX type of toxin by characterization of the aqxCABD operon. All hemolytic A. equuli isolates contained a functional aqxCABD operon and expressed the Aqx hemolysin as shown by genetic and phenotypic assays. The structural toxin AqxA is the hemolysin of A. equuli as shown by expression of recombinant aqx constructs in E. coli. Its hemolytic activity can be inhibited by specific antibodies raised against AqxA. Sequence analysis of the 16S rRNA gene (rrs) of the taxonomically diffuse group of A. equuli and related strains defined two phylogenetically distinct groups. The presence of the Aqx operon is not correlated with this phylogenetic grouping. The operon is found in both groups of A. equuli strains where it specifies the hemolytic activity and is supposedly to be a determinative virulence factor. The aqx operon was not found in closely related members of the Pasteurellaceae family. The description of the Aqx hemolysin will open new ways for studying the pathogenesis of A. equuli.

Analysis of non-porcine isolates of Actinobacillus suis

Twenty-four Actinobacillus suis isolates obtained from several species of non-porcine mammals were compared to the representative porcine strains, ATCC 15557 (serotype O1) and H89-1173 (serotype O2), by O serotyping, DNA fingerprinting, PCR amplification of apxICA, apxIICA and apxIIICA toxin genes and by rrs (16S rRNA) gene sequencing. Only two strains, both equine, reacted with O1 antiserum while two others, one canine and the other feline, reacted with O2 antiserum. One equine strain reacted weakly with both antisera. No amplification of apx genes was found with the non-porcine O1 or the "not O1/O2" strains but amplification of the apxICA and apxIICA genes was observed with the two O2 strains. In addition, these two O2 strains had both BamHI and BglII fingerprints that were very similar to the porcine O2 reference strain, H89-1173 and rrs gene sequences that were identical to the A. suis reference strain ATCC 15557. Taken together, these data suggest that although many non-porcine A. suis isolates are not A. suis (sensu stricto), some isolates are genotypically as well as phenotypically similar to A. suis.

Proposal of a new serovar of Actinobacillus pleuropneumoniae: serovar 15

We report on the re-examination of nine Australian isolates of Actinobacillus pleuropneumoniae that have been previously assigned to serovar 12. In the ring precipitation test, none of the nine isolates reacted with antisera to serovars 1-14 of A. pleuropneumoniae. Antiserum prepared against one of the Australian isolates gave no reaction with any of the 14 recognised serovar reference strains, except serovar 7. This reaction of the HS143 antiserum with serovar 7 antigen could be removed by adsorption with serovar 7 antigen. The adsorbed antiserum remained reactive with HS143 and the other eight Australian isolates. The nine Australian isolates were all shown to express ApxII and ApxIII, found in serovars 2, 4, 6 and 8, as well as the 42kDa outer membrane protein found in all serovars of A. pleuropneumoniae. The nine Australian isolates were found to possess the following toxin associated genes apxIBD, apxIICA, apxIIICA, apxIIIBD and apxIVA. The toxin gene profile of the Australian isolates is typical of A. pleuropneumoniae serovars 2, 4, 6 and 8. On the basis of the serological characterisation results and the toxin gene profiles, we propose that these isolates represent a new serovar of A. pleuropneumoniae--serovar 15--with HS143 being the reference strain for the new serovar.

Identification and detection of Actinobacillus pleuropneumoniae by PCR based on the gene apxIVA

The apxIVA gene, a recently discovered RTX determinant of Actinobacillus pleuropneumoniae, was shown to be species-specific. DNA hybridization experiments using probes for various regions of apxIVA revealed that the 3'-terminus of this gene was present in all 14 serotypes of A. pleuropneumoniae but absent from phylogenetically related species. A primer pair spanning this region specifically amplified a 422bp fragment in PCR experiments with DNA from the reference strains of the 14 serotypes and 194 field strains isolated from various geographic locations worldwide. DNA sequence analysis of PCR products derived from all serotypes were identical except in serotypes 3, 8, and 10, which showed minor differences. The PCR did not amplify any product when DNA from 17 different bacterial species closely related to A. pleuropneumoniae was used as template. In addition, the PCR was negative with DNA of several Actinobacillus sp. which were initially characterized as A. pleuropneumoniae using routine phenotypic and serological analyses but which were subsequently shown by 16S rRNA sequence analysis to belong to yet undefined Actinobacillus species. The sensitivity of the PCR was determined to be 10pg of A. pleuropneumoniae DNA. A set of nested primers amplified a 377bp fragment specifically with A. pleuropneumoniae DNA. DNA titration experiments using the flanking and nested primer pairs showed an improved level of sensitivity to approximately 10fg of genomic DNA. The nested PCR was used to monitor the spread of A. pleuropneumoniae in pigs experimentally infected with a virulent serotype 1 strain and housed in a controlled environment facility. A. pleuropneumoniae DNA could be detected by nested PCR in nasal swab samples of infected pigs receiving either a high dose (5x10(5)) or a low dose (1x10(4)) challenge and in unchallenged cohorts that were contact-infected by the inoculated animals. Furthermore, PCR confirmed the presence of A. pleuropneumoniae in 16/17 homogenates from necrotic lung lesions, while the bacterium was successfully recovered from 13 of these lesions by culture.

Characterization of apxIVA, a new RTX determinant of Actinobacillus pleuropneumoniae

A fourth type of RTX determinant was identified in Actinobacillus pleuropneumoniae and was designated apxIVA. When expressed in Escherichia coli, recombinant ApxIVA showed a weak haemolytic activity and co-haemolytic synergy with the sphingomyelinase (beta-toxin) of Staphylococcus aureus. These activities required the presence of an additional gene, ORF1, that is located immediately upstream of apxIVA. The apxIVA gene product could not be detected in A. pleuropneumoniae cultures grown under various conditions in vitro; however, pigs experimentally infected with A. pleuropneumoniae serotypes 1, 5 and 7 started to produce antibodies that reacted with recombinant ApxIVA 14 d post-infection, indicating that apxIVA is expressed in vivo. In addition, sera from pigs naturally and experimentally infected with any of the serotypes all reacted with recombinant ApxIVA. The apxIVA gene from the serotype 1 A. pleuropneumoniae type strain Shope 4074T encodes a protein with a predicted molecular mass of 202 kDa which has typical features of RTX proteins including hydrophobic domains in the N-terminal half and 24 glycine-rich nonapeptides in the C-terminal half that bind Ca2+. The glycine-rich nonapeptides are arranged in a modular structure and there is some variability in the number of modules in the ApxIVA proteins of different serotypes of A. pleuropneumoniae. The deduced amino acid sequences of the ApxIVA proteins have significant similarity with the Neisseria meningitidis iron-regulated RTX proteins FrpA and FrpC, and to a much lesser extent with other RTX proteins. The apxIVA gene could be detected in all A. pleuropneumoniae serotypes and seems to be species-specific. Although the precise role of this new RTX determinant in pathogenesis of porcine pleuropneumonia needs to be determined, apxIVA is the first in vivo induced toxin gene that has been described in A. pleuropneumoniae.

Frequency of patient-reported infections among sicker adults in high-income countries: an international perspective

The frequency of patient-reported health care-associated infections across several high-income countries was analyzed in representative population samples based on data from "The Commonwealth Fund's 2011 International Survey of Sicker Adults in Eleven countries." Across countries, 8.9% of patients who were hospitalized and/or had surgery reported an infection, but this rate varied considerably from 5.3% in the United States to 11.9% in New Zealand. Patients who reported infection were more likely to rate the quality of medical care received as fair or poor (odds ratio [OR], 2.4; 95% confidence interval [CI]: 1.9-3.1, P < .001). Female sex (OR, 1.2; 95% CI: 1.0-1.5, P = .027), reporting 2 or more chronic conditions (OR, 1.5; 95% CI: 1.1-2.0, P = .004), poor health (OR, 1.6; 95% CI: 1.2-2.1, P < .001), and surgery (OR, 1.8; 95% CI: 1.4-2.3, P < .001) were significant predictors for health care-associated infection across countries. Being above 64 years of age (OR, 0.78; 95% CI: 0.64-0.95, P = .013) and day-surgery (OR, 0.62; 95% CI: 0.48-0.79, P < .001) decreased the likelihood for reporting infection.

Valuing the Health States Associated with Chlamydia trachomatis Infections and Their Sequelae: A Systematic Review of Economic Evaluations and Primary Studies

OBJECTIVES Economic evaluations of interventions to prevent and control sexually transmitted infections such as Chlamydia trachomatis are increasingly required to present their outcomes in terms of quality-adjusted life-years using preference-based measurements of relevant health states. The objectives of this study were to critically evaluate how published cost-effectiveness studies have conceptualized and valued health states associated with chlamydia and to examine the primary evidence available to inform health state utility values (HSUVs). METHODS A systematic review was conducted, with searches of six electronic databases up to December 2012. Data on study characteristics, methods, and main results were extracted by using a standard template. RESULTS Nineteen economic evaluations of relevant interventions were included. Individual studies considered different health states and assigned different values and durations. Eleven studies cited the same source for HSUVs. Only five primary studies valued relevant health states. The methods and viewpoints adopted varied, and different values for health states were generated. CONCLUSIONS Limitations in the information available about HSUVs associated with chlamydia and its complications have implications for the robustness of economic evaluations in this area. None of the primary studies could be used without reservation to inform cost-effectiveness analyses in the United Kingdom. Future debate should consider appropriate methods for valuing health states for infectious diseases, because recommended approaches may not be suitable. Unless we adequately tackle the challenges associated with measuring and valuing health-related quality of life for patients with chlamydia and other infectious diseases, evaluating the cost-effectiveness of interventions in this area will remain problematic.

Effectiveness and cost-effectiveness of traditional and new partner notification technologies for curable sexually transmitted infections: observational study, systematic reviews and mathematical modelling

BACKGROUND Partner notification is essential to the comprehensive case management of sexually transmitted infections. Systematic reviews and mathematical modelling can be used to synthesise information about the effects of new interventions to enhance the outcomes of partner notification. OBJECTIVE To study the effectiveness and cost-effectiveness of traditional and new partner notification technologies for curable sexually transmitted infections (STIs). DESIGN Secondary data analysis of clinical audit data; systematic reviews of randomised controlled trials (MEDLINE, EMBASE and Cochrane Central Register of Controlled Trials) published from 1 January 1966 to 31 August 2012 and of studies of health-related quality of life (HRQL) [MEDLINE, EMBASE, ISI Web of Knowledge, NHS Economic Evaluation Database (NHS EED), Database of Abstracts of Reviews of Effects (DARE) and Health Technology Assessment (HTA)] published from 1 January 1980 to 31 December 2011; static models of clinical effectiveness and cost-effectiveness; and dynamic modelling studies to improve parameter estimation and examine effectiveness. SETTING General population and genitourinary medicine clinic attenders. PARTICIPANTS Heterosexual women and men. INTERVENTIONS Traditional partner notification by patient or provider referral, and new partner notification by expedited partner therapy (EPT) or its UK equivalent, accelerated partner therapy (APT). MAIN OUTCOME MEASURES Population prevalence; index case reinfection; and partners treated per index case. RESULTS Enhanced partner therapy reduced reinfection in index cases with curable STIs more than simple patient referral [risk ratio (RR) 0.71; 95% confidence interval (CI) 0.56 to 0.89]. There are no randomised trials of APT. The median number of partners treated for chlamydia per index case in UK clinics was 0.60. The number of partners needed to treat to interrupt transmission of chlamydia was lower for casual than for regular partners. In dynamic model simulations, > 10% of partners are chlamydia positive with look-back periods of up to 18 months. In the presence of a chlamydia screening programme that reduces population prevalence, treatment of current partners achieves most of the additional reduction in prevalence attributable to partner notification. Dynamic model simulations show that cotesting and treatment for chlamydia and gonorrhoea reduce the prevalence of both STIs. APT has a limited additional effect on prevalence but reduces the rate of index case reinfection. Published quality-adjusted life-year (QALY) weights were of insufficient quality to be used in a cost-effectiveness study of partner notification in this project. Using an intermediate outcome of cost per infection diagnosed, doubling the efficacy of partner notification from 0.4 to 0.8 partners treated per index case was more cost-effective than increasing chlamydia screening coverage. CONCLUSIONS There is evidence to support the improved clinical effectiveness of EPT in reducing index case reinfection. In a general heterosexual population, partner notification identifies new infected cases but the impact on chlamydia prevalence is limited. Partner notification to notify casual partners might have a greater impact than for regular partners in genitourinary clinic populations. Recommendations for future research are (1) to conduct randomised controlled trials using biological outcomes of the effectiveness of APT and of methods to increase testing for human immunodeficiency virus (HIV) and STIs after APT; (2) collection of HRQL data should be a priority to determine QALYs associated with the sequelae of curable STIs; and (3) standardised parameter sets for curable STIs should be developed for mathematical models of STI transmission that are used for policy-making. FUNDING The National Institute for Health Research Health Technology Assessment programme.

Strategies for partner notification for sexually transmitted infections, including HIV

BACKGROUND Partner notification (PN) is the process whereby sexual partners of an index patient are informed of their exposure to a sexually transmitted infection (STI) and the need to obtain treatment. For the person (index patient) with a curable STI, PN aims to eradicate infection and prevent re-infection. For sexual partners, PN aims to identify and treat undiagnosed STIs. At the level of sexual networks and populations, the aim of PN is to interrupt chains of STI transmission. For people with viral STI, PN aims to identify undiagnosed infections, which can facilitate access for their sexual partners to treatment and help prevent transmission. OBJECTIVES To assess the effects of different PN strategies in people with STI, including human immunodeficiency virus (HIV) infection. SEARCH METHODS We searched electronic databases (the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE and EMBASE) without language restrictions. We scanned reference lists of potential studies and previous reviews and contacted experts in the field. We searched three trial registries. We conducted the most recent search on 31 August 2012. SELECTION CRITERIA Published or unpublished randomised controlled trials (RCTs) or quasi-RCTs comparing two or more PN strategies. Four main PN strategies were included: patient referral, expedited partner therapy, provider referral and contract referral. Patient referral means that the patient notifies their sexual partners, either with (enhanced patient referral) or without (simple patient referral) additional verbal or written support. In expedited partner therapy, the patient delivers medication or a prescription for medication to their partner(s) without the need for a medical examination of the partner. In provider referral, health service personnel notify the partners. In contract referral, the index patient is encouraged to notify partner, with the understanding that the partners will be contacted if they do not visit the health service by a certain date. DATA COLLECTION AND ANALYSIS We analysed data according to paired partner referral strategies. We organised the comparisons first according to four main PN strategies (1. enhanced patient referral, 2. expedited partner therapy, 3. contract referral, 4. provider referral). We compared each main strategy with simple patient referral and then with each other, if trials were available. For continuous outcome measures, we calculated the mean difference (MD) with 95% confidence intervals (CI). For dichotomous variables, we calculated the risk ratio (RR) with 95% CI. We performed meta-analyses where appropriate. We performed a sensitivity analysis for the primary outcome re-infection rate of the index patient by excluding studies with attrition of greater than 20%. Two review authors independently assessed the risk of bias and extracted data. We contacted study authors for additional information. MAIN RESULTS We included 26 trials (17,578 participants, 9015 women and 8563 men). Five trials were conducted in developing countries. Only two trials were conducted among HIV-positive patients. There was potential for selection bias, owing to the methods of allocation used and of performance bias, owing to the lack of blinding in most included studies. Seven trials had attrition of greater than 20%, increasing the risk of bias.The review found moderate-quality evidence that expedited partner therapy is better than simple patient referral for preventing re-infection of index patients when combining trials of STIs that caused urethritis or cervicitis (6 trials; RR 0.71, 95% CI 0.56 to 0.89, I(2) = 39%). When studies with attrition greater than 20% were excluded, the effect of expedited partner therapy was attenuated (2 trials; RR 0.8, 95% CI 0.62 to 1.04, I(2) = 0%). In trials restricted to index patients with chlamydia, the effect was attenuated (2 trials; RR 0.90, 95% CI 0.60 to 1.35, I(2) = 22%). Expedited partner therapy also increased the number of partners treated per index patient (three trials) when compared with simple patient referral in people with chlamydia or gonorrhoea (MD 0.43, 95% CI 0.28 to 0.58) or trichomonas (MD 0.51, 95% CI 0.35 to 0.67), and people with any STI syndrome (MD 0.5, 95% CI 0.34 to 0.67). Expedited partner therapy was not superior to enhanced patient referral in preventing re-infection (3 trials; RR 0.96, 95% CI 0.60 to 1.53, I(2) = 33%, low-quality evidence). Home sampling kits for partners (four trials) did not result in lower rates of re-infection in the index case (measured in one trial), or higher numbers of partners elicited (three trials), notified (two trials) or treated (one trial) when compared with simple patient referral. There was no consistent evidence for the relative effects of provider, contract or other patient referral methods. In one trial among men with non-gonococcal urethritis, more partners were treated with provider referral than with simple patient referral (MD 0.5, 95% CI 0.37 to 0.63). In one study among people with syphilis, contract referral elicited treatment of more partners than provider referral (MD 2.2, 95% CI 1.95 to 2.45), but the number of partners receiving treatment was the same in both groups. Where measured, there was no statistical evidence of differences in the incidence of adverse effects between PN strategies. AUTHORS' CONCLUSIONS The evidence assessed in this review does not identify a single optimal strategy for PN for any particular STI. When combining trials of STI causing urethritis or cervicitis, expedited partner therapy was more successful than simple patient referral for preventing re-infection of the index patient but was not superior to enhanced patient referral. Expedited partner therapy interventions should include all components that were part of the trial intervention package. There was insufficient evidence to determine the most effective components of an enhanced patient referral strategy. There are too few trials to allow consistent conclusions about the relative effects of provider, contract or other patient referral methods for different STIs. More high-quality RCTs of PN strategies for HIV and syphilis, using biological outcomes, are needed.

Sonication: a valuable technique for diagnosis and treatment of periprosthetic joint infections

BACKGROUND Periprosthetic joint infection (PJI) is the most severe complication, following joint arthroplasty. Identification of the causal microbial factor is of paramount importance for the successful treatment. PURPOSE The aim of this study is to compare the sonication fluid cultures derived from joint prosthetic components with the respective periprosthetic tissue cultures. METHODS Explanted prosthesis components for suspected infection were placed into a tank containing sterile Ringer's solution and sonicated for 1 minute at 40 kHz. Sonication fluid cultures were examined for 10 days, and the number and identity of any colony morphology was recorded. In addition, periprosthetic tissue specimens (>5) were collected and cultured according to standard practice. The duration of antimicrobial interruption interval before culture sampling was recorded. RESULTS Thirty-four patients composed the study group. Sonication fluid cultures were positive in 24 patients (70.5%). Sixteen of thirty four periprosthetic tissue cultures (47.1%) were considered positive, all revealing the same microbial species with the respective sonication fluid cultures: 3 tissue samples showed polymicrobial infection. All tissue cultures were also found positive by the sonication fluid culture. CONCLUSIONS Sonication fluid cultures represent a cheap, easy, accurate, and sensitive diagnostic modality demonstrating increased sensitivity compared to periprosthetic tissue cultures (70.5 versus 47.1%).

Catheter removal versus retention in the management of catheter-associated enterococcal bloodstream infections

BACKGROUND Enterococci are an important cause of central venous catheter (CVC)-associated bloodstream infections (CA-BSI). It is unclear whether CVC removal is necessary to successfully manage enterococcal CA-BSI. METHODS A 12-month retrospective cohort study of adults with enterococcal CA-BSI was conducted at a tertiary care hospital; clinical, microbiological and outcome data were collected. RESULTS A total of 111 patients had an enterococcal CA-BSI. The median age was 58.2 years (range 21 to 94 years). There were 45 (40.5%) infections caused by Entercoccus faecalis (among which 10 [22%] were vancomycin resistant), 61 (55%) by Enterococcus faecium (57 [93%] vancomycin resistant) and five (4.5%) by other Enterococcus species. Patients were treated with linezolid (n=51 [46%]), vancomycin (n=37 [33%]), daptomycin (n=11 [10%]), ampicillin (n=2 [2%]) or quinupristin/dalfopristin (n=2 [2%]); seven (n=6%) patients did not receive adequate enterococcal treatment. Additionally, 24 (22%) patients received adjunctive gentamicin treatment. The CVC was retained in 29 (26.1%) patients. Patients with removed CVCs showed lower rates of in-hospital mortality (15 [18.3%] versus 11 [37.9]; P=0.03), but similar rates of recurrent bacteremia (nine [11.0%] versus two (7.0%); P=0.7) and a similar post-BSI length of hospital stay (median days [range]) (11.1 [1.7 to 63.1 days] versus 9.3 [1.9 to 31.8 days]; P=0.3). Catheter retention was an independent predictor of mortality (OR 3.34 [95% CI 1.21 to 9.26]). CONCLUSIONS To the authors' knowledge, the present article describes the largest enterococcal CA-BSI series to date. Mortality was increased among patients who had their catheter retained. Additional prospective studies are necessary to determine the optimal management of enterococcal CA-BSI.