Improvement in Neisseria gonorrhoeae culture rates by bedside inoculation and incubation at a clinic for sexually transmitted infections.

BACKGROUND: Culture of Neisseria gonorrhoeae is essential for surveillance of complete antimicrobial susceptibility profiles. In 2014, the culture success rate of N. gonorrhoeae from samples taken at the clinic for sexually transmitted infections (STI clinic), Oslo University Hospital, Norway, was only 20%. The present study aimed to improve gonococcal culture rates using bedside inoculation of patient samples on gonococcal agar plates and incubation at the STI clinic. METHODS: This prospective quality improvement study was conducted by the STI clinic and the Department of Microbiology at Oslo University Hospital from May 2016 - October 2017. When culture of N. gonorrhoeae was clinically indicated, we introduced a parallel 'bedside culture' at the STI clinic and compared results with the standard culture at the microbiology department. Samples were taken from urethra, anorectum, pharynx and cervix. Culture rates were compared across symptomatic and asymptomatic anatomical sites. RESULTS: From 596 gonococcal-positive PCR samples, bedside culture had a significantly higher success rate of 57% compared to 41% with standard culture (p < 0.05). Overall, culture rate from symptomatic sites was 91% v. 45% from asymptomatic sites. The culture rates from different anatomical sites were as follows: urethra 93%, anorectum 64%, pharynx 28% and cervix 70%. Bedside culture significantly (p < 0.05) improved the culture rates for symptomatic urethral and asymptomatic pharyngeal samples. CONCLUSIONS: Where feasible, bedside inoculation on gonococcal agar plates and incubation of samples from patients with gonorrhoea is recommended. This will improve the culture diagnostics and provide additional gonococcal isolates for antimicrobial resistance surveillance.

Epidemiology and spa-type diversity of meticillin-resistant Staphylococcus aureus in community and healthcare settings in Norway.

BACKGROUND: There has been a marked increase in the incidence of meticillin-resistant Staphylococcus aureus (MRSA) during the past decade in Norway; a country with one of the lowest prevalence rates and an active 'search-and-destroy' policy applied to hospital settings. AIM: To characterize the trends of notification rates of community-associated (CA) and healthcare-associated (HA) MRSA in Norway, and explore the diversity and circulation of MRSA spa types within and outside healthcare settings. METHODS: A registry-based study on notified MRSA infections and colonizations was conducted in Norway between 2006 and 2015. The diversity and abundance of CA- and HA-MRSA spa types were compared using novel ecological diversity measures (Hill numbers). FINDINGS: During the study period, the monthly notification rate increased 6.9-fold and 1.8-fold among CA- and HA-MRSA, respectively; the increase was steeper among colonizations than infections. In both settings, the distribution of spa types was uneven, with a few dominant spa types and many singletons. The spa-type diversity of CA-MRSA was higher than HA-MRSA in terms of different types (685 vs 481), and increased during the study period. However, the diversity associated with the dominant spa types was similar and remained stable. A high overlap of spa types was estimated between the settings; spa-t002, t019 and t008 were the most common. CONCLUSION: The present findings suggest a strong connection between CA- and HA-MRSA epidemiology in Norway. If the fast-growing trend of CA-MRSA continues in the years to come, it may challenge current guidelines and infection control of MRSA in healthcare environments.

High frequency of occupied attB regions in Norwegian Staphylococcus aureus isolates supports a two-step MRSA screening algorithm.

Rapid nucleic acid amplification tests for methicillin-resistant Staphylococcus aureus (MRSA) diagnostics commonly target the mec resistance gene, genes specific for S. aureus, and the integration site for the SCCmec resistance cassette, orfX. Due to poor specificity when these target genes are used individually, additional culture is required to verify positive results. The combination of these targets is useful, but the optimal algorithm may depend on the presence of the genetic markers in S. aureus isolates, as well as the prevalence of MRSA in a population. The aim of the present study was to identify a rapid, low-cost, and functional screening algorithm in order to reduce the response time for MRSA diagnostics. An in-house orfX-SCCmec polymerase chain reaction (PCR) assay was established and evaluated. The results were compared with an existing mec/nuc PCR assay and traditional culture. Methicillin-sensitive S. aureus (MSSA) that tested false-positive in the orfX-SCCmec PCR assay were further investigated with full genome sequencing using the Ion PGM™ System to verify results and causality. Based on these data, a two-step screening algorithm with initial mec/nuc PCR followed by orfX-SCCmec PCR on positive samples was suggested and tested on 1443 patient samples. 22.5 % of MSSA isolates tested false-positive with the orfX-SCCmec PCR. Full genome sequencing of these isolates identified genetic variation in the attB region of S. aureus, including empty cassette variants and non-mec SCC. The suggested two-step MRSA screening algorithm allowed us to report MRSA results for 95.6 % of all samples and 99 % of MRSA-negative samples after one day.

DNA microarray analysis of Staphylococcus aureus causing bloodstream infection: bacterial genes associated with mortality?

Providing evidence for microbial genetic determinants' impact on outcome in Staphylococcus aureus bloodstream infections (SABSI) is challenging due to the complex and dynamic microbe-host interaction. Our recent population-based prospective study reported an association between the S. aureus clonal complex (CC) 30 genotype and mortality in SABSI patients. This follow-up investigation aimed to examine the genetic profiles of the SABSI isolates and test the hypothesis that specific genetic characteristics in S. aureus are associated with mortality. SABSI isolates (n = 305) and S. aureus CC30 isolates from asymptomatic nasal carriers (n = 38) were characterised by DNA microarray analysis and spa typing. Fisher's exact test, least absolute shrinkage and selection operator (LASSO) and elastic net regressions were performed to discern within four groups defined by patient outcome and characteristics. No specific S. aureus genetic determinants were found to be associated with mortality in SABSI patients. By applying LASSO and elastic net regressions, we found evidence suggesting that agrIII and cna were positively and setC (=selX) and seh were negatively associated with S. aureus CC30 versus non-CC30 isolates. The genes chp and sak, encoding immune evasion molecules, were found in higher frequencies in CC30 SABSI isolates compared to CC30 carrier isolates, indicating a higher virulence potential. In conclusion, no specific S. aureus genes were found to be associated with mortality by DNA microarray analysis and state-of-the-art statistical analyses. The next natural step is to test the hypothesis in larger samples with higher resolution methods, like whole genome sequencing.

Population-based epidemiology of Staphylococcus aureus bloodstream infection: clonal complex 30 genotype is associated with mortality.

Staphylococcus aureus bloodstream infections (SABSI) are associated with a high burden of morbidity and mortality. The impact of specific S. aureus genotypes on outcome is unclear. The aim of this study was to evaluate the epidemiology and outcome of SABSI, with a special emphasis on the impact of bacterial clonal lineage on mortality. We conducted a 3-year population-based prospective study between 2011 and 2014, including 303 consecutive adult patients. Clinical data were obtained from interviews and medical records. S. aureus isolates were genotyped using DNA microarrays. The incidence rate of SABSI was 27.6 per 100,000 inhabitants [95 % confidence interval (CI) 24.6-31.0]. The median age of the patients was 71 years (interquartile range 56-81 years) and 61.4 % were male. Most SABSI (70.6 %) occurred in hospitals or associated to healthcare, and 34.1 % of these were associated with intravascular catheters. Only five (1.6 %) SABSI were caused by methicillin-resistant S. aureus (MRSA). The 30-day case fatality rate was 20.8 % (95 % CI 16.6-25.7). S. aureus clonal complex 30 [hazard ratio (HR) 3.9; 95 % CI 1.8-8.5, p = 0.001], unknown focus of infection (HR 4.5; 95 % CI 1.9-10.8, p = 0.001) and respiratory tract infection (HR 12.7; 95 % CI 4.6-34.6, p < 0.001) were independent predictors of mortality in a Cox regression analysis after adjusting for age, sex and underlying conditions. A high proportion of potential preventable SABSI calls for effective infection control measures. S. aureus clonal complex 30 genotype was associated with mortality in patients with bloodstream infections. The genetic basis underlying this association remains to be demonstrated.

Molecular characterization of VIM-producing Klebsiella pneumoniae from Scandinavia reveals genetic relatedness with international clonal complexes encoding transferable multidrug resistance.

VIM-producing Klebsiella pneumoniae (VPKP) has been identified as a source of hospital outbreaks and is prevalent particularly in the Mediterranean region. In this study we have characterized eight VPKP isolates identified in Scandinavia during 2005-2008. With the exception of one isolate, all were from patients with recent history of hospitalization abroad (Greece, n = 6; Turkey, n = 1). Multilocus sequence typing (MLST) resulted in five sequence types (STs), ST36 (n = 1), ST147 (n = 4), ST272 (n = 1), ST273 (n = 1) and ST383 (n = 1), which except for ST272 were part of putative international clonal complexes. All were multidrug resistant due to the presence of other resistance determinants, including extended-spectrum ß-lactamases (CTX-M-3, SHV-5 and SHV-12), 16S rRNA methylases (ArmA) and plasmid-mediated quinolone resistance determinants (QnrS). One isolate harboured a novel VIM-variant (VIM-26) while VIM-1 and VIM-19 were detected in six and one isolate, respectively. Two different genetic structures surrounding the bla(VIM) gene were identified in four isolates. In two isolates bla(VIM-1) and bla(VIM-26) were located in an integron similar to In-e541 (intI1;bla(VIM-1/-26);aacA7; dhfrI;aadA1;3'CS) while in the other two isolates bla(VIM-1) was located in an integron lacking 3'CS but with an IS26 element in the 3'end (intI1;bla(VIM-1);aac(6')-Ib;IS26), as identified in the IncN plasmid pKOX105. The bla(VIM) -genes were located on transferable plasmids ranging from ∼40 to ∼240 kb and associated with Tn21 in four isolates. PCR-based replicon typing indicated association of bla(VIM) with IncN (n = 3) and A/C (n = 1) broad-host-range plasmids but also with unknown replicons (n = 4). In conclusion, Scandinavian VPKP is associated with importation and genetically related to international clones encoding transferable plasmid-mediated multidrug resistance.

Do HIV-seropositive patients become colonised with drug-resistant microorganisms?

The aim of the present study was to investigate whether HIV-infected patients, a group that is supposedly at risk for infection with antibiotic-resistant microbes, really does so, and to assess possible risk factors for acquiring these organisms. During the period from January 1998 to July 1999, samples of normal flora were obtained from 107 HIV-infected patients attending an outpatient clinic in Oslo, Norway. The samples were cultured for Streptococcus pneumoniae, Haemophilus influenzae, Escherichia coli, coagulase-negative staphylococci and Candida spp., and the resulting isolates were tested for antimicrobial susceptibility. The patients studied represented all stages of HIV infection, from recently infected to severely immunocompromised. Samples were taken at one, two or three time-points to determine whether antimicrobial resistance in colonising microorganisms increases over time. Antimicrobial resistance was linked primarily to antimicrobial prophylaxis, but it did not increase during the observation period. The level of a patient's immunodeficiency and the consequently intensified medical care was also of some importance. Even though about 50% of the patients were receiving antimicrobial agents at the time of sampling, the level of resistance found in these patients was very similar to that found in other patient groups in Norway; except for Candida albicans isolates, which were less susceptible to fluconazole. Overall, antimicrobial resistance was uncommon in the HIV-seropositive patients studied, a finding that is probably related to the overall low prevalence of antimicrobial resistance in the general population in Norway.

Antibiotic sensitivity still prevails in Norwegian blood culture isolates.

We describe the antimicrobial susceptibility of bacteraemia isolates from Norway. From March 1998 to February 1999, four university hospitals covering all parts of Norway collected their first 10 isolates each month. Minimal inhibitory concentrations were determined for: Enterobacteriaceae (n=192), staphylococci (n=89) and Streptococcus pneumoniae (n=69) using the Etest. NCCLS breakpoints were used. About 20% of all blood culture isolates in Norway in this period were investigated. Compared with countries outside Scandinavia antibiotic sensitivity still prevails. Only minor differences in resistance were found between participating hospitals, between hospital departments and between hospital- and community-acquired pathogens. The prudent use of antibiotics in Norway may contribute to the fact that antibiotic resistance still remains low in the most common bacterial pathogens causing bloodstream infections.

Emerging antibiotic resistance in Salmonella typhimurium in Norway.

The antimicrobial resistance of 809 Salmonella Typhimurium isolates collected from humans in Norway between 1975 and 1998 was studied. The material was subdivided into domestic and foreign isolates according to whether the patient had recently travelled abroad or not. In imported isolates the largest increase in resistance was in 1996 when 35% of the isolates were multi-resistant. The first multi-resistant isolate acquired in Norway appeared in 1994, but already in 1998 23% of the isolates domestically acquired were multi-resistant, and a majority were S. Typhimurium DT104. We found no ciprofloxacin resistance in domestically acquired isolates. Amplified fragment length polymorphism analysis was performed on selected multi-resistant isolates. The method discriminated well between different multi-resistant isolates, but not between DT104 isolates. Resistant and multi-resistant S. Typhimurium were until 1998 essentially recovered from patients who had travelled abroad, but multi-resistant isolates, mainly DT104, are now also being transmitted within the country.

Low occurrence of antibiotic resistance in Escherichia coli and staphylococci isolated from blood cultures in two Norwegian hospitals in 1991-92 and 1995-96.

The aim of this study was to investigate the antibiotic resistance rates of major bacterial pathogens causing bloodstream infections in two very different types of hospital in Norway. We examined all Escherichia coli and staphylococci (330 isolates) causing bloodstream infections from one general county hospital and one specialist national cancer hospital during the periods 1991-92 and 1995-96. Minimal inhibitory concentrations (MICs) were determined using the E-test. E. coli and staphylococci constituted 46.7% of all isolates from bloodstream infections in the two hospitals. Overall, E. coli isolates were resistant to amoxicillin (21%), trimethoprim (21%), doxycycline (20%) and trimethoprim-sulphamethoxazole (17%), while Staphylococcus aureus strains were resistant to benzylpenicillin (66%). No methicillin-resistant S. aureus was detected. Coagulase-negative staphylococci were often multiresistant, but remained fully sensitive to vancomycin. For a few antibiotics, significantly more resistance was found in the specialist hospital. In our material we found no significant increase in resistance between 1991-92 and 1995-96. In conclusion, antimicrobial resistance still remains low in important bacterial pathogens causing bloodstream infections in Norway.

Antibiotic resistance mechanisms in Salmonella species causing bacteraemia in Malawi and Kenya.

In two studies on the causative agents of bacteraemia in Malawi and Kenya, 33 Salmonella strains were isolated. Fourteen strains of Salmonella typhimurium and Salmonella enteritidis were found to exhibit resistance to amoxicillin, amoxicillin/clavulanic acid and cotrimoxazole as well as decreased susceptibility to a range of aminoglycosides. The resistant strains were studied to establish their resistance mechanisms. Beta-lactamase co-focusing with TEM-1 was present in 12 strains. In two strains, both S. typhimurium from Kenya, an OXA-1 beta-lactamase was detected. The aminoglycoside-modifying enzyme ANT(2") was found in 10 strains. The presence of the encoding genes was confirmed by PCR. For comparison, susceptibility records of 73 Salmonella strains isolated during the past 14 years in our hospital were studied retrospectively. Only one of these strains was resistant to amoxicillin. This resistance was acquired during therapy.