- Volume 66, Issue 6, 2017

Purpose. The standard approach to screening sexually transmitted infections (STIs) has often been restricted to urogenital specimens. Most current guidelines, however, also recommend testing extra-genital sites, including rectal locations, because asymptomatic rectal carriage of pathogens has often been reported. The aim of our study was to evaluate self-collected rectal swabs to screen bacterial STIs in HIV-infected patients in Marseille, France.

Methodology. Between January 2014 and December 2015, 118 HIV-infected patients (93 males and 25 females) agreed to self-sample anal swabs for detection of bacterial STI. Detection of Neisseria gonorrhoeae, Chlamydia trachomatis, Treponema pallidum, Mycoplasma genitalium and Haemophilus ducreyi was performed using in-house qPCR assay.

Results/Key findings. Bacterial STIs were found in 8 % (9/118) of the patients. C. trachomatis was the most commonly detected bacterium (4.2 %) followed by N. gonorrhoeae (2.5 %), M. genitalium (1.7 %) and T. pallidum (0.8 %). All the positive patients were males. The rectal carriage of pathogenic bacteria was fortuitously discovered for seven men (78 %) who did not present rectal signs of STIs and was suspected for two men who presented proctitis (22 %).

Conclusion. In conclusion, testing extra-genital sites is crucial for the diagnosis of STIs in men and women presenting or not concomitant urogenital infections in order to detect asymptomatic carriage with the aim of controlling and preventing transmission to their sexual partners.

Purpose. A study was undertaken to determine the risk factors and trends in antimicrobial resistance for enteric fever.

Methodology. Demographic, antimicrobial susceptibility, typing and epidemiological data were examined for 2005–2012 in patients with enteric fever in London. Single and multivariable logistic regression was used to determine the risk factors associated with antibiotic resistance.

Results. 453 cases with Salmonella enterica subsp. enterica serovar Paratyphi A, 17 with S. Paratyphi B and 611 with S. enterica subsp. enterica serovar Typhi were examined. For travellers, 335 (88 %) of S. Paratyphi A isolates were resistant to ciprofloxacin, but resistance to other antimicrobials was low. Almost 80 % (395) of the S. Typhi isolates were resistant to ciprofloxacin, 131 (26 %) to ampicillin, 131 (27 %) to chloramphenicol, 137 (28 %) to trimethoprim and 171 (28 %) to sulphonamide. None of the isolates were resistant to cephalosporins.

A trend analysis for S. Typhi isolates showed no significant change in resistance to ampicillin, chloramphenicol, sulphonamide and trimethoprim or for multidrug resistance (P=0.38). Overall resistance to ciprofloxacin increased for S. Paratyphi A (P=0.018) and for S. Typhi (P<0.001) but fell for S. Typhi in 2011–2012. Resistance profiles were reflected by specific phage types and countries visited by the travellers.

Conclusions. The proportion of S. Typhi strains resistant to ampicillin, chloramphenicol and cotrimoxazole remained steady for the period 2005–2012. There was a significant increase in a trend for resistance to ciprofloxacin which increased until 2010, followed by a fall in 2011–2012. S. Paratyphi resistance to ciprofloxacin increased until 2012. Specific phage types were associated with resistance to specific antimicrobials and travel abroad.

Purpose. Under certain circumstances, Actinomyces behaves as an opportunistic microorganism and can cause actinomycosis, a chronic and inflammatory granulomatous infection. The purpose of this project was to detect the presence of Actinomyces in cervical exudates from women with cervical intraepithelial neoplasia (CIN) and women with cervical cancer.

Methodology. Cervical samples from 92 women were divided into three groups: CIN, cervical cancer and healthy women. Metagenomic DNA extraction was performed following the Qiagen QIAamp Mini Kit protocol. A specific fragment (675 bp) was amplified by PCR in order to detect the presence of Actinomycetales. Samples in which Actinomycetales was detected were subjected to separate amplification reactions with primer pairs for A. israelii, A. viscosus, A. meyeri and A. odontolyticus. Amplified products were observed by 2 % agarose gel electrophoresis.

Results. Actinomyces were found in 10 % of women with CIN, 36.6 % of women with cervical cancer and 9 % of healthy women. The species identified in this study were A. meyeri in 14/92 samples (15.2 %), A. viscosus in 10/92 samples (10.8 %), A. odontolyticus in 4/92 samples (4.3 %) and A. israelii in 6/92 samples (6.5 %).

Conclusion. Patients with cervical cancer had a higher prevalence of the presence of Actinomyces compared to the CIN and control groups. This is the first study in which a deliberate search of this genus has been performed in women with cervical pathologies. The use of specific primers for each species facilitated their detection in comparison with traditional isolation methods. More information is necessary to understand the molecular mechanisms involved in the complex role that bacterial communities may play in the development of cancer (and vice versa).

Purpose. The aim of this study was to compare the in vitro activity of ampicillin and moxifloxacin against six isolates selected from 154 invasive clinical isolates of Listeria monocytogenes and evaluate their intra- and extracellular activities with achievable central nervous system concentrations obtained using Monte Carlo simulations with conventional and unconventional dosages.

Methodology. The MICs and minimal bactericidal concentrations (MBCs) of ampicillin and moxifloxacin were determined by using the broth microdilution method. The intra- and extracellular activities were compared using time–kill curves and inhibition of intracellular growth assays.

Results. The MICs50/90 of ampicillin were 0.125/0.5 mg l−1 and the MBC50/90 was ≥16 mg l−1, while the moxifloxacin MICs50/90 were 0.25/0.5 mg l−1 and the MBC50/90 was 0.5 mg l−1. Ampicillin did not show any extracellular bactericidal activity at 24 h, although bactericidal activity was detected at 48 h. For moxifloxacin, the bactericidal effect was evident after 6 h of incubation. Both antibiotics achieved significant reductions in intracellular inoculum after 1–24 h of incubation; however, moxifloxacin becomes bactericidal more rapidly, producing a much greater reduction in the inoculum in the first hour than ampicillin. There were no differences among the MIC and MBC values of moxifloxacin and ampicillin among the strains belonging to different serotypes and/or epidemic clones. This fact was also found in the intra- and extracellular studies.

Conclusion. The results of this study demonstrated the faster bactericidal activity of moxifloxacin at achievable central nervous system concentrations against intra- and extracellular forms of L. monocytogenes in comparison with ampicillin.

Purpose. Methicillin-resistant Staphylococcus aureus (MRSA) colonizes the skin of hospitalized patients and is associated with high morbidity and mortality. To prevent colonization and infection by S. aureus, better disinfection practices are required. Therefore, we evaluated the effect of chlorhexidine whole-body washing on hospital-acquired S. aureus infections among intensive care unit (ICU) patients in a tertiary hospital in Mexico.

Methodology. The study was conducted over 18 months to evaluate the effect of 2 % chlorhexidine gluconate (CXG) whole-body washing of ICU adult patients on chlorhexidine and antibiotic resistance, biofilm production and clonal distribution of S. aureus in a tertiary care hospital. Minimum inhibitory concentrations for CXG, antibiotic susceptibility and biofilm production by S. aureus isolates were determined. Pulsed-field gel electrophoresis, multilocus sequence typing (MLST) and PCR for Panton–Valentine leucocidin (PVL) were used for molecular typing of MRSA isolates.

Results/Key findings. We included 158 isolates. A reduction in antibiotic resistance in the study period was observed for clindamycin, levofloxacin, norfloxacin, oxacillin and trimethoprim/sulfamethoxazole. None of the isolates showed reduced susceptibility to CXG. Most of the isolates were non-biofilm producers (147/158). The most commonly identified clone was a descendant of the ST5-MRSA-II (New York/Japan) clone. This clone decreased during the intervention period and reappeared markedly in the post-intervention period. During the post-intervention period, two isolates were related with the clone ST8-MRSA-IV (also known as USA300).

Conclusion. Our findings suggest that the CXG bathing favored the reduction of healthcare-associated MRSA isolates and a temporary reduction of the predominant ST5-MRSA-II (New York/Japan) clone.

Purpose. An observational study was performed to investigate the carriage rate and serotypes of Streptococcus pneumoniae in the 13-valent pneumococcal conjugate vaccine (PCV13) era in Taiwan.

Methodology. From March 2014 to March 2015 a total of 500 healthy children and their households (631 adults) were enrolled from two large medical centres for nasopharyngeal carriage survey. Clinical isolates were prospectively collected from June 2012 to May 2015 at Chang Gung Memorial Hospital. We applied a multiplex polymerase chain reaction in addition to culture to detect S. pneumoniae.

Results. S. pneumoniae was isolated from 12.0 % of the children and 3.6 % of the households. In the children's cohort only 23.3 % of the isolates could be assigned to PCV13 serotypes; non-vaccine serotypes were predominant (76.6 %) and the most frequently detected non-vaccine serotypes were 15A/F and 15B/C (both 13.3 %), followed by 23A (6.7 %). In the household cohort, 21.7 % belonged to PCV13 serotypes, and 78.3 % to non-vaccine serotypes. Clinical analysis of culture-confirmed pneumococcal infection showed that infection caused by PCV13 serotypes decreased by 47 % from 83 % in 2012–2013 to 44 % in 2014–2015, while infection caused by non-PCV13 serotypes increased from 17 to 56 %. Among the carriage isolates a significantly higher percentage belonged to serogroup 15 compared to serogroup 19 (26.6 vs 6.66 %, 2014–2015; P=0.003). Therefore, clinical isolates belonging to serogroup 15 were more prevalent than those belonging to serogroup 19 (44.1 vs 32.3 %, 2014–2015; P=0.318).

Conclusion. The isolation of non-vaccine serotypes and unknown serotypes after the introduction of PCV13 in children highlights the importance of continued surveillance for emerging serotypes.

Purpose. To examine whether the epidemiology of bacteraemia caused by methicillin-susceptible Staphylococcus aureus (MSSA) and methicillin-resistant S. aureus (MRSA) differed in children aged <1 year and in comparison to older age groups.

Methodology. English mandatory MRSA and MSSA surveillance data from 2006 and 2011, respectively, were collected. Epidemiological information was descriptively analysed in relation to methicillin susceptibility and patient age. Ninety-five percent confidence intervals (CIs) are reported.

Results/Key findings. The average incidence rate of MSSA and MRSA bacteraemia in <1-year-olds was 60.2 and 4.8 episodes per 100 000 population per year, respectively. Of the cases of MSSA bacteraemia in children aged <1 year, 47.5 % (95 % CI: 45.1–50.0; n=760/1 599) were in neonates. With increasing age up to one year, more MSSA bacteraemias were detected ≥7 days after admission, ranging from 0 % (95 % CI: 0–2.5 %) in 0–2-day-olds to 68.4 % (95 % CI: 64.0–72.5 %; 333/487) in 8–28-day-olds and 50.5 % (95 % CI: 47.1–54.0 %; 423/837) in 29 day–1-year-olds, a higher proportion than in older children but similar to MRSA bacteraemia. Amongst <1-year-olds with MSSA bacteraemia, the underlying source was most commonly recorded as intravascular devices [34.4 % (95 %, CI: 30.5–38.6 %); n=190/552] whilst in older age groups this declined. A similar trend was observed for MRSA bacteraemia.

Conclusions. Our analysis indicates that S. aureus bacteraemia in <1-year-olds is primarily healthcare-associated, unlike MSSA bacteraemia in older age groups. Paediatric-specific interventions targeted at the healthcare setting, such as neonatal unit-specific care bundles and paediatric device-specific strategies, are required.

Purpose. Catheter-related bloodstream infections (CRBSIs) are among the most common hospital-acquired infections. We aimed to survey methicillin resistance, biofilm production and susceptibility to vancomycin, linezolid and other antibiotics for staphylococci isolated from CRBSIs.

Methodology. Fifty-eight isolates [20 S . aureus and 38 coagulase-negative staphylococci (CoNS; 20 Staphylococcus epidermidis, nine Staphylococcus haemolyticus, three Staphylococcus schleiferi, two Staphylococcus warneri and four Staphylococcus lugdunensis)] were tested for methicillin resistance by cefoxitin disk diffusion and detection of the mecA gene by PCR; biofilm-forming ability using Congo red agar and tissue culture plate methods; susceptibility to ciprofloxacin, clindamycin, cotrimoxazole, erythromycin, gentamicin, linezolid, rifampicin and tetracycline; and MIC determination for vancomycin.

Results/Key findings. Cefoxitin resistance was detected among 40 % (8/20) S . aureus isolates, 70 % (14/20) S . epidermidis isolates and 16.7 % (3/18) of other CoNS, although the mecA gene was detected in 45 % (9/20) S . aureus isolates, 35 % (7/20) S . epidermidis isolates and 16.7 % (3/18) of other CoNS. Biofilm-forming ability ranged from 45 to 75 %. Methicillin-resistant S. aureus and other CoNS were considered to be more virulent than methicillin-resistant S. epidermidis due to the higher biofilm forming abilities of the former. All tested isolates exhibited 100 % sensitivity to vancomycin and linezolid, irrespective of their methicillin resistance or biofilm-forming ability. Rifampicin showed overall sensitivity of 75.9 %. Varying degrees of multi-resistance were found for the other antibiotics.

Conclusion. Vancomycin, linezolid and rifampicin could be used effectively against methicillin-resistant staphylococci isolated from CRBSIs.

Purpose. The role of antibiotics below their MIC in the development of bacterial drug resistance is becoming increasingly important. We investigated the effect of sub-MICs of bactericidal antibiotics on the susceptibility pattern of Staphylococcus aureus and evaluated the role of free radicals.

Methodology. A total of 12 S. aureus strains were recovered from pus samples and their antibiograms determined. The test isolates were treated with sub-MIC levels of tetracycline, gentamicin, ciprofloxacin and cefotaxime. Alterations in their respective breakpoints were observed along with measurements of free radical generation by nitro blue tetrazolium test.

Results/Key findings. Gentamicin, ciprofloxacin and cefotaxime exposure significantly altered the breakpoints of exposed isolates against several tested antibiotics and higher levels of free radicals were generated after antibiotic exposure.

Conclusions. Our study demonstrates that sub-MIC levels of antimicrobials can lead to resistance and cross-resistance across several classes of antibiotics in wild strains of S. aureus, possibly by free radical production. The molecular mechanisms behind the acquisition of drug resistance at low antibiotic concentrations and the specific target genes of reactive oxygen speciesneed to be explored further.

Purpose. Sitafloxacin (SFX) is a new fluoroquinolone (FQ) that has shown a strong bactericidal effect against Mycobacterium tuberculosis (Mtb) in vitro. However, data on SFX efficacy against Mtb with gyrA/B mutations and its epidemiological cut-off (ECOFF) value remain limited. Therefore, we evaluated and compared the in vitro activity of SFX against gyrA/B-mutant Mtb to that of moxifloxacin (MFX), levofloxacin (LFX) and ciprofloxacin (CFX), and determined the ECOFF for SFX.

Methodology. A total of 109 clinical Mtb isolates, including 73 multidrug-resistant (MDR) isolates, were subjected to minimum inhibitory concentration (MIC) analysis in oleic-albumin-dextrose-catalase (OADC)-supplemented Middlebrook 7H9 medium. Our results showed that SFX had lower cumulative MIC than MFX, LFX and CFX. Furthermore, we performed direct DNA sequencing of the quinolone-resistance-determining regions (QRDRs).

Results. We identified the following mutations: D94G, D94A, A90V, D94H, D94N and G88A in gyrA; and A543V, A543T, E540D, R485C, D500A, I552S and D577A in gyrB. Based on our results, an ECOFF of 0.125 µg ml−1 was proposed for SFX. With this ECOFF, 15 % of LFX-resistant isolates with MIC ≥2 µg ml−1 were susceptible to SFX.

Conclusion. SFX had the lowest cumulative MIC and a relatively low ECOFF value against Mtb, indicating that SFX was not only more effective against gyrA-mutant isolates, but also MDR isolates in Japan.

Purpose. The purpose of this study was to evaluate the usefulness of a new diagnostic multiplexed bead-based bioassay (Quantamatrix Multiplexed Assay Platform; QMAP) system with shape-encoded silica microparticles for the rapid and accurate detection and identification of 23 mycobacterial species/groups, including Mycobacterium tuberculosis complex (MTBC).

Methodology. A total of 295 mycobacterial clinical isolates cultured from respiratory specimens were used for identification of MTBC and non-tuberculous mycobacteria (NTM) using the QMAP system and the results were confirmed with PCR-restriction fragment length polymorphism (RFLP) analysis of the rpoB gene, rpoB sequence analysis and PCR-reverse blot hybridization assay (REBA).

Results/Key findings. The Mycobacterium genus-specific probe of the QMAP system was positive for all 46 Mycobacterium reference strains and negative for 59 non-Mycobacterium strains. Based on 295 liquid culture-positive samples, both the culture-based conventional identification method and the QMAP system identified each 212 and 81 isolates as MTB and NTM species. The concordance rates for the identification of NTM species between the QMAP system and molecular assays were 92.8 % (77/83), 97.6 % (81/83) and 100 % (83/83) for PCR-RFLP, the rpoB sequence analysis and PCR-REBA, respectively.

Conclusion. The QMAP system yielded rapid, highly sensitive and specific results for the identification of MTBC and NTM and accurately discriminated between NTM species within 3 h.

Purpose. Surveillance of the bacterial spectrum and antibiotic-resistance patterns of locally occurring uropathogens is essential to serve as a basis for empirical treatment of urinary tract infections (UTIs), as antibiotic-resistance rates may vary geographically with significant differences between countries and regions, and with time.

Methodology. We retrospectively analysed all urine samples taken in the department of urology in a tertiary care hospital in Hungary from January 2004 to December 2015.

Results/Key findings. The five most commonly occurring bacteria were Escherichia coli, Enterococcus faecalis, Klebsiella pneumoniae, Pseudomonas aeruginosa and Proteus mirabilis. Resistance of Escherichia coli to ciprofloxacin increased significantly from 19 to 25 %. Although the resistance of Escherichia coli against cephalosporins showed an increasing trend, it still remained generally low. However, resistance rates of K. pneumoniae to cephalosporins were very high, reaching 60 %, due to the high rate of extended-spectrum-β-lactamase-positive Klebsiella strains. We observed a significant increase in the rate of carbapenem-resistant Pseudomonas aeruginosa.

Conclusion. Fluoroquinolones cannot be recommended for empirical treatment in our region. Cephalosporins can be a good empirical choice for treating Gram-negative UTIs, but should be avoided when multi-drug resistant (MDR) bacteria are suspected. Increases in the rate of carbapenem-resistant Pseudomonas aeruginosa, and in the general rate of MDR bacteria, are both a very alarming trend. We recommend practising prudent antibiotic policy, preferably using antibiotics with the narrowest possible spectrum.

Purpose. Diabetic patients seem to be predisposed to cutaneous candidiasis. In this study, we evaluated the interference of diabetic conditions in alloxan-induced diabetic mice in relation to the development of C. albicans infection, density of M1 and M2 macrophages, distribution of collagen type I and III and anti-inflamamatory cytokines involved in tissue repair.

Methodology. The mice were treated with intravenous alloxan, and all animals with blood glucose levels >250  mg dl−1 were inoculate with C. albicans intradermally in the hind paw and were studied for up to 21 days. Control groups without alloxan were used. The fungal burden was evaluated by periodic acid-Schiff (PAS) and by counting the colony forming units. Total population of macrophages were targeted with antibody to F4/80 antigen and M2 macrophages with anti-arginase antibody. Anti-inflammatory cytokines from popliteal lymph nodes were determined by capture ELISA procedures. Picrosirius red staining allowed qunantification of collagen types I and III in the infected skin by using a polarized light microscope.

Results/Key findings. Diabetic mice, versus non-diabetic mice, showed a significant lower density of F4/80 and M2 macrophages, higher fungal burden, deficiency in interleukin (IL)-4 production, and delayed IL-13 responses. The later clearance of C. albicans enhanced tissue injury, leading to a decrease in collagen type I. Moreover, collagen type III was increased by interference of IL-13 and transforming growth factor-β cytokines.

Conclusion. These findings highlight some important changes in diabetic animal responses to C. albicans infection that may be important to the pathophysiological processes underpinning cutaneous candidiasis in diabetic patients.

Purpose. Staphylococcus aureus infections have contributed to the global healthcare burden, particularly with regard to hospital-acquired meticillin-resistant S. aureus (MRSA) infections.

Methodology. This study describes the antibacterial activity of diacetylcurcumin (DAC) against meticillin-susceptible S. aureus/MRSA biofilm formation, survival, metabolic activity and structure; its ability to prevent bacterial adhesion to human cells; and toxicity in Galleria mellonella larvae.

Results. DAC showed excellent antibacterial activity, with MIC ranging between 17.3 and 34.6 µmol l−1, and minimum bactericidal concentration ranging between 69 and 277 µmol l−1. It significantly reduced bacterial biofilm survival – by 22–63 % (at MIC, 10×MIC or 100×MIC) as compared to the 25–42 % reduction by vancomycin (P<0.0001) – and severely affected biofilm cell structures, leading to damaged architecture and the formation of amorphous cell clusters. Treatment with DAC (MIC/4) decreased bacterial adhesion to HaCaT keratinocytes from 1 to 5 h (P<0.0001). Finally, DAC demonstrated low toxicity in G. mellonella at its effective anti-biofilm concentrations.

Conclusion. These findings open new avenues for the study of this curcumin derivative as an excellent prototype with anti-MRSA activity.

Purpose. Prevotella spp. represent a diverse genus of bacteria, frequently identified by both culture and molecular methods in the lungs of patients with chronic respiratory infection. However, their role in the pathogenesis of chronic lung infection is unclear; therefore, a more complete understanding of their molecular epidemiology is required.

Methodology. Pulsed Field Gel Electrophoresis (PFGE) and Random Amplified Polymorphic DNA (RAPD) assays were developed and used to determine the degree of similarity between sequential isolates (n=42) from cystic fibrosis (CF) patients during periods of clinical stability and exacerbation.

Results. A wide diversity of PFGE and RAPD banding patterns were observed, demonstrating considerable within-genus heterogeneity. In 8/12 (66.7 %) cases, where the same species was identified at sequential time points, pre- and post-antibiotic treatment of an exacerbation, PFGE/RAPD profiles were highly similar or identical. Congruence was observed between PFGE and RAPD (adjusted Rand coefficient, 0.200; adjusted Wallace RAPD->PFGE 0.459, PFGE->RAPD 0.128). Furthermore, some isolates could not be adequately assigned a species name on the basis of 16S rRNA analysis: these isolates had identical PFGE/RAPD profiles to Prevotella histicola.

Conclusion. The similarity in PFGE and RAPD banding patterns observed in sequential CF Prevotella isolates may be indicative of the persistence of this genus in the CF lung. Further work is required to determine the clinical significance of this finding, and to more accurately distinguish differences in pathogenicity between species.