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Abstract Clinical isolates (n = 63) of Pseudomonas aeruginosa obtained from various sites in 63 horses were compared using ERIC2 RAPD PCR to determine their genetic relatedness. Resulting banding patterns (n = 24 genotypes) showed a high degree of genetic heterogeneity amongst all isolates examined, indicating a relative non-clonal relationship between isolates from these patients, employing this genotyping technique. This study characterised 63 clinical isolates into 24 distinct genotypes, with the largest cluster (genotype E) accounting for 10/63 (15.9%) of the isolates. ERIC2 RAPD PCR proved to be a highly discriminatory molecular typing tool of P. aeruginosa in isolates recovered from horses. With the adoption of several controls to aid reproducibility, this technique may be useful as an alternative to PFGE, particularly in epidemiological investigations of outbreaks where speed may be a significant parameter. This is the first report of clonal heterogeneity amongst P. aeruginosa from horses and demonstrated that ERIC RAPD PCR is a rapid method for the examination of this species in horses, which may be useful in outbreak analysis.

The Gram-negative bacterium, Pseudomonas aeruginosa , is a major respiratory pathogen in patients with cystic fibrosis (CF), with an associated increase in morbidity and mortality. Consequently, infection prevention and control (IPC) plays an important role within health care in order to minimize the risk of cross-infection of this organism amongst patients and the hospital environment. It was the aim of this study to examine bacterial contamination of the health estate of CF in-patients' single-bedded rooms and related environments (n=40). Twelve bacterial genera were identified, six being Gram-positive (Brevibacterium, Dermacoccus, Micrococcus, Rothia, Staphylococcus and Streptococcus), and six being Gram-negative ( Acinetobacter, Citrobacter, Klebsiella, Moraxella, Pantoea and Pseudoxanthomonas). None of the organisms identified were considered of particular clinical significance to CF patients. The CF lung and associated sputa may be important reservoirs of Pseudomonas aeruginosa, with potential for spill-over into the health care estate. In the aftermath of the Pseudomonas neonatal outbreak at Altnagelvin and the Royal Jubilee Maternity Hospitals, where there was heightened IPC awareness regarding the presence of this bacterium, it is encouraging to note its absence from the CF-health care estate examined., Competing Interests: Provenance: externally peer reviewed., (Copyright © 2020 Ulster Medical Society.)

Competing Interests: Provenance: externally peer reviewed

Competing Interests: Provenance: externally peer reviewed

Competing Interests: Provenance: externally peer reviewed

Competing Interests: Provenance: externally peer reviewed

The aim of this study was to examine the microbiological and related parameters (antibiotic resistance and pathogen identification) of water at two salmonid fish farms in Northern Ireland. Total Bacterial Counts at the Movanagher Fish Farm was 1730 colony forming units (cfu)/ml water (log10 3.24cfu/ml) and 3260cfu/ml (log10 3.51cfu/ml) at the Bushmills Salmon Station. Examination of resulting organisms revealed 10 morphological phenotypes, which were subsequently sequenced to determine their identification. All these organisms were Gram-negative and no Gram-positive organisms were isolated from any water sample. From these phenotypes, eight different genera were identified including Acinetobacter, Aeromonas, Chryseobacterium, Erwinia, Flavobacterium, Pseudomonas and Rheinheimera. One unnamed novel taxon was identified from water at the Movanagher Fish Farm, belonging to the genus Acinetobacter and has been tentatively named Acinetobacter movanagherensis. No other novel taxa were observed. All but one of these environmental organisms (Erwinia) are potential pathogens of fish disease. Total antibiotic resistance was observed to varying degrees in water specimens. The most resistant populations were observed in water taken from the Bushmills Salmon Station inlet, followed by water from the Movanagher Fish Farm. No resistance was observed against tetracycline and there was only one occurrence of resistance against ciprofloxacin. Overall, this study indicates that potential fish pathogens made up the majority of environmental organisms identified, even in the absence of recorded fish disease. There was also relatively high levels of total antibiotic resistance in the bacterial water populations examined, where tetracycline was the only antibiotic with zero resistance. These data indicate that the threat of bacterial disease is relatively close due to the indigenous colonization of farm water and that husbandry standards should be maintained at a high standard to avert bacterial disease outbreaks, rather than relying on the absence of specific pathogens in the immediate farm environment., (Copyright © 2014 Elsevier Inc. All rights reserved.)

A study was undertaken to examine the population structure of viridans group streptococci (VGS) isolated the upper respiratory tract of adult and paediatric patients within the community. VGS are common commensal bacterial inhabitants of the upper respiratory tract and valuable sentinel reporters of underlying antibiotic resistance (AR). Laboratory examination of the colonising VGS species may provide a valuable ecological description of the species isolated from the upper respiratory tract and their antibiotic susceptibility, including an estimation of the AR reservoir in this population. Freshly obtained nasal and oropharyngeal swabs from 84 patients were examined by selective conventional culture on Mitis-Salivarius agar and yielded 363 isolates of VGS. Sequence analyses of the rpnB and 16-23S rRNA ITS genes identified these isolates to belong to 10 species of VGS and included S. anginosus, S. australis, S. constellatus, S. infantis, S. mitis, S. oralis, S. parasanguinis, S. salivarius, S. sanguinis and S. vestibularis. The most frequent VGS organisms isolated was S. salivarius (282/363; 78.0%), followed by S. sanguinis (23/363; 6.3%), S. parasanguinis (21/363; 5.8%), S. mitis (18/363; 5.0%), S. anginosus (5/363; 1.4%), S. vestibularis (5/363; 1.4%), S. australis (3/363; 0.8%), S. oralis (3/363; 0.8%), S. infantis (1/363; 0.3%) and S. constellatus (1/363; 0.3%). All patients examined carried at least one VGS organism, where there were 17 combination patterns of carriage of the 10 species of VGS species isolated, where 54.2%, 37.3%, 7.2% and 1.2% of patients harboured one, two, three and four different VGS species, respectively. Antibiotic susceptibility was determined by standard disk diffusion assay testing against four classes of antibiotics, including the b-lactams [cefotaxime, cefuroxime], the tetracyclines [doxycycline], the fluoroquinolones [levofloxacin] and the macrolides [erythromycin]. Overall, there was no resistance to levofloxacin and cefuroxime, with limited resistance to cefotaxime (3.3%) and doxycycline (9.8%). Antibiotic resistance was highest in erythromycin, where 40.9% of isolates were resistant. S. vestibularis was the most antibiotic resistance of all VGS species examined (S. vestibularis v S. salivarius p=0.011), followed by S. anginosis. S. salivarius was the most antibiotic susceptible VGS species examined. Overall, given their infrequency in causing infection, relatively few studies to date have attempted to examine their ecology in their preferred body niche, namely the upper respiratory tract. However, knowing their prevalence is becoming increasingly important in relation to their ability to exclude significant respiratory pathogens, including Streptococcus pneumoniae. In conclusion, these data indicate that VGS colonisation of the upper respiratory tract in individuals within the community is dominated mainly with relatively antibiotic susceptible S. salivarius.

Antibiotic resistance in clinical pathogens in humans may be traced back to resistance mechanisms in environmental bacteria and any factors, which are likely to alter (upregulate) resistance in environmental organisms, is of potential and eventual consequence to human pathogens. Furthermore, sublethal doses of gamma radiation to environmental organisms may cause sublethal stress and a selective pressure, which may lead to mutational events that alter the bacterium's susceptibility profile. A gamma (γ) radiation simulation experiment was performed to emulate the exposure of four environmental bacteria, including Listeria innocua, Bacillus subtilis, E. coli and Pseudomonas aeruginosa, to levels of radiation in and around Fukushima, Japan, equating to 1, 10 and 100 years equivalence exposure. Alteration to susceptibility to 14 antibiotics was measured as the primary endpoint. There was no significant alteration in the susceptibility of the Gram-positive organisms, whereas both Gram-negative organisms became slightly more susceptible to the antibiotics tested over time. These data indicate that such radiation exposure will not increase the antibiotic resistance profile of these organisms and hence not add to the global public health burden of increased antibiotic resistance in human bacterial pathogens., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Streptococcus pneumoniae is the leading cause of community-acquired pneumonia (CAP). Currently, empirical treatment with quinolones is being used due to the emergence of beta-lactam and macrolide resistance in S. pneumonaie. Although the prevalence of quinolone-resistant S. pneumoniae remains low, increasing numbers of resistant isolates are being seen. Genetic mechanisms leading to fluoroquinolone resistance in pneumococci are complex. This study aims to use molecular methods to characterise all isolates through sequence analysis of their QRDR regions. Thirty-two S. pneumoniae isolates were obtained from nasal swabs from adult and paediatric patients attending local general practices in Northern Ireland. Phenotypic minimum inhibitory concentration (MIC) was determined for Clinical and Laboratory Standards Institute (CLSI) broth microdilution against ciprofloxacin, levofloxacin and norfloxacin. Simultaneously, the QRDR regions of gyrA, gyrB, parC and parE were analysed by sequence typing for all pneumococci obtained. Only one isolate (3.1%) showed reduced susceptibility to ciprofloxacin and levofloxacin. Two amino acid positions were discordant in the S. pneumoniae R6 strain and eight (25%) and 23 (71.9%) isolates contained the mutations Ile460Val in gyrA and Lys137Asn in parC (deposited in GenBank, accession numbers GQ999587-GQ999589), respectively. No mutations were found in either the gyrB or parE loci. In conclusion, the study demonstrated increased fluoroquinolone resistance which could not be accounted for simply through QRDR mutations, and, reciprocally, that mutations in the QRDR region do not necessarily result in overt phenotypic resistance.

Inadvertent exposure of bacterial pathogens to X-ray radiation may be an environmental stress, where the bacterium may respond by increasing mutational events, thereby potentially resulting in increased antibiotic resistance and alteration to genotypic profile. In order to examine this, four clinical pathogens, including the Gram-negative organisms Escherichia coli O157:H7 NCTC12900 and Pseudomonas aeruginosa NCTC10662, as well as the Gram-positive organisms Staphylococcus aureus NCTC6571 and Enterococcus faecium were exposed to X-rays (35,495 cGy/cm2) over a seven-day period. Antibiotic susceptibility was assessed before, during and after exposure by examining susceptibility, as quantified by E-test with six antibiotics, as well as to a further 11 antibiotics by measurement of susceptibility zone sizes (mm). Additionally, the DNA profile of each organism was compared before, during and after exposure employing the enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC PCR). Results indicated that exposure of these organisms to this amount of X-ray radiation did not alter their antibiotic susceptibility, nor their genomic DNA profile. Overall, these data indicate that exposure of bacteria to X-ray radiation does not alter the test organisms' antibiotic susceptibility profiles, nor alter genomic DNA profiles of bacteria, which therefore does not compromise molecular epidemiological tracking of bacteria within healthcare environments in which patients have been exposed to X-ray radiation.

Although about 75-80% of neutropenic fevers are thought to be caused by infections, a causal organism can be confirmed microbiologically or suspected clinically in only 30-50%, and even fewer of these cases (16%) have a documented bacteraemia. The cause of neutropenic fever in the remaining cases remains elusive. The reasons for this failure may be due to the difficulty in recovering low numbers of organisms, fastidious organisms which fail to grow using conventional culture media, the presence of non-culturable organisms, or the presence of inhibitory substances in specimens. Previously, the authors showed the presence of Acinetobacter in peripheral blood of febrile neutropenic patients with a haematological malignancy, using 16S rDNA polymerase chain reaction (PCR) and sequencing techniques. However, conventional culture was unable to detect these organisms. Hence, it was felt necessary to examine the antibacterial properties of four antineoplastic agents used in the treatment of haematological malignancy, namely bleomycin, cisplatin, doxorubicin and vincristine. A total of 56 wild-type Acinetobacter including seven species (A. calcoaceticus [n=17], A. septicus [n=11], A. baumannii [n=10], A. johnsonii [n=7], A. lwoffii [n=8] A. haemolyticus [n=2] and A. radioresistens [n=1]) were examined for their susceptibility to the four antineoplastic agents at therapeutic concentration. No inhibition was observed, but inhibition was seen at higher concentrations of both bleomycin and doxorubicin. Time to detection of blood culture bottles containing separate antineoplastic agents (i.e., bleomycin and doxorubicin) was compared to that containing saline using a paired t-test. Samples containing doxorubicin at 1 pg/mL were shown to have a mean time to detection of 21.8 h (range: 15.6-31.4 h). Bottles containing saline had a mean time to detection of 22.9 h (range: 18.2-31.3 h). Statistical analysis showed no significant difference (P=0.3361) between time to detection for blood culture bottles containing doxorubicin at achievable plasma concentration and corresponding negative controls. With regard to bleomycin (300 miu/mL), the mean time to detection was 27.29 h (range: 20.2-38.4 h) in the test bottles, with mean time to detection in the saline negative controls of 22.56 h (range: 17.0-30.1 h). Paired t-test gave P=0.000451, hence a significant difference in time to detection for blood cultures containing therapeutic levels of bleomycin. Overall, the antineoplastic agents vincristine, cisplatin or doxorubicin did not have any inhibitory effects on the Acinetobacter organisms examined. At worst, therapeutic concentrations of bleomycin may delay automated detection of an Acinetobacter bacteraemia by a mean time of 5.9 h.

The aim of this study was to investigate the reliability of disc diffusion testing with penicillin, erythromycin and ciprofloxacin within the viridans group streptococci (VGS). In total, the antibiotic susceptibilities of 167 VGS isolates were compared by standard disc diffusion and broth microdilution methods, and these phenotypic data were compared to the carriage of the respective gene resistance determinants [ermB and mefA/E (macrolides); QRDR, gyrA, gyrB, parC and parE (quinolones)]. Overall, there were 35 discrepancies [resistant by MIC and susceptible by zone diameter (21.0%)] between MIC and disc diameter when penicillin susceptibility was interpreted by Clinical and Laboratory Standards Institute criteria. Scattergrams showed a bimodal distribution between non-susceptible and susceptible strains when erythromycin susceptibility was tested by both methods. Thirty-four (20.4%) isolates were categorized as resistant by MIC breakpoints, while disc diameter defined these as having intermediate resistance. With ciprofloxacin, three isolates (1.8%) showed minor discrepancies between MIC breakpoints and disc diameter. Isolates non-susceptible to all three antimicrobial agents tested were reliably distinguished from susceptible isolates by disc diffusion testing, except for the detection of low-level resistance to penicillin, where broth microdilution or an alternative quantitative MIC method should be used. Otherwise, we conclude that disc diffusion testing is a reliable method to detect strains of VGS non-susceptible to penicillin, erythromycin and ciprofloxacin, as demonstrated with their concordance to their gene resistance characteristics., (© 2011 SGM)

Breakthrough contamination of tuberculosis (TB) cultures is a problem in that it allows the overgrowth of another bacterium present in the sputum specimen, which can potentially mask the presence of Mycobacterium tuberculosis. The aim of this study was to isolate and characterize the bacterial organisms responsible for such overgrowth and contamination, and to examine their susceptibility to (i) various chemical selective decontamination steps and (ii) antibiotics in liquid culture media, in an attempt to develop a method to help alleviate contamination problems associated with the conventional isolation of M. tuberculosis from routine patient sputum specimens. Bacterial contaminants from 102 routine sputum cultures were identified molecularly by 16S rRNA gene PCR and direct sequencing from contaminated Löwenstein-Jensen (LJ) slopes and BacT/Alert liquid medium. It was found that the contaminants from LJ slopes belonged to 11 different genera and were composed largely of Gram-negative organisms (84.9 %; 45/53), whereas the liquid culture contaminants belonged to 13 different genera, with 37/66 isolates (56.1 %) being Gram-negative. Pseudomonas aeruginosa was the dominant contaminant in both media. The effect of six different selective decontamination protocols was examined. Four of the six methods were effective at eliminating all culturable organisms present; these were 5 % oxalic acid, 5 % oxalic acid/2 % NaOH, 5 % oxalic acid/4 % NaOH and 1 % chlorhexidine. NaOH at a concentration of 2 or 4 % was less effective as it was unable to eliminate all organisms of each species tested, with the exception of P. aeruginosa. In conclusion, breakthrough contamination of TB cultures is due to a diverse range of at least 17 different bacterial genera, with P. aeruginosa and Staphylococcus epidermidis accounting for the dominant contaminating flora. Employment of chemical decontaminating protocols solely involving NaOH may lead to higher rates of contamination. Where such contamination is encountered, TB laboratories should consider the reprocessing of such sputum samples with an alternative decontamination method such as 1 % chlorhexidine.

Purpose: To describe the clinical outcomes of a compulsory switch from branded to generic levetiracetam (LEV) among people with epilepsy (PWE) in an outpatient setting., Methods: We conducted a retrospective chart review of 760 unduplicated consecutive adult patients attending a tertiary care epilepsy clinic at Ben Taub General Hospital. On November 1, 2008 hospital policy required all patients receiving branded LEV to be automatically switched to generic LEV. We calculated the proportion of patients switching back to branded LEV and reasons for the switch back., Key Findings: Of the 260 patients (34%) being prescribed LEV (generic and brand name) during the study period, 105 (42.9%) were switched back to brand name LEV by their treating physicians. Reasons for switch back included increase in seizure frequency (19.6% vs. 1.6%; p < 0.0001) and adverse effects (AEs) (3.3%). AEs included headache, fatigue, and aggression. Patient age was associated with switchback when controlling for gender, epilepsy classification, and treatment characteristics [relative risk (RR) 2.44; 95% confidence interval (CI) 2.09-2.84; p < 0.05)]. An increase in seizure frequency subsequent to generic substitution was associated with polytherapy compared to monotherapy (3.225; 1.512-6.880; p < 0.05)., Significance: A significant proportion of patients in our cohort on generic LEV required switch back to the branded drug. Careful monitoring is imperative because a compulsory switch from branded to generic LEV may lead to poor clinical outcomes, with risk of AEs and increased seizure frequency., (Wiley Periodicals, Inc. © 2011 International League Against Epilepsy.)

A study was undertaken to examine the population structure of viridans group streptococci (VGS) in the sputum of adult patients with cystic fibrosis (CF). Freshly expectorated sputa (n=58) from 45 adult CF patients were examined by selective conventional culture on Mitis-Salivarius agar and yielded 190 isolates of VGS. Sequence analyses of the rpnB and 16-23S rRNA ITS genes identified these isolates to belong to 12 species of VGS and included S. anginosus, S. australis, S. cristatus, S. gordonii, S. infantis, S. mitis, S. mutans, S. oralis, S. parasanguinis, S. pneumoniae, S. salivarius and S. sanguinis. The most frequently VGS organism isolated was S. salivarius (47/190; 24.7%), followed by S. mitis (36/190; 19%), S. sanguinis (25/190; 13.2%), S. oralis (20/190; 11.0%), S. pneumoniae (19/190; 10.0%), S. parasanguinis (16/190; 8.4%), S. infantis (11/190; 5.8%), S. gordonii (7/190; 3.7%), S. anginosus (4/190; 2.1%), S. cristatus (2/190; 1.1%), S. australis (1/190; 0.5%), S. mutans (1/190; 0.5%) and S. agalactiae (1/190; 0.5%). All, but four, patients harboured at least one VGS species, which ranged from one to five streptococcal species, with a mean of 2.85 species per patient. There was no clonality at the subspecies level employing ERIC RAPD PCR. Antibiotic susceptibility was determined by Minimum Inhibitory Concentration (MIC) testing against penicillin, erythromycin and ciprofloxacin. Overall, resistance to penicillin with all VGS was 73/190 (38.4%) and 167/190 (87.9%) for erythromycin. With regard to ciprofloxacin, 27/190 (14.2%) were fully resistant, whilst a further 21/190 (11.1%) showed intermediate resistance, which equated to approximately three quarters (74.7%) of isolates being fully sensitive to this agent. In addition, as a comparator control population, we examined antibiotic susceptibility, as above, in a non-CF population comprising 12 individuals (50 VGS isolates), who were not receiving chronic antibiotics. In comparison, 8% and 38% of VGS isolates from non-CF individuals were resistant by disk susceptibility testing to penicillin and erythromycin, respectively. None of the non-CF VGS organisms were resistant to ciprofloxacin, but 42% showed intermediate resistance., (Copyright © 2010 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.)

Viridans-group streptococci (VGS) consist of several taxa which historically have been highly diverse. However, at times it may become necessary to have a reliable scheme for the identification of these organisms to the species level. The aim of this study is to compare the ability of five gene loci, namely rnpB, 16S rRNA, 16S-23S rRNA, sodA and dnaJ, to speciate such organisms through a sequence typing-based approach. Reference organisms consisting of six VGS species were compared based on sequence typing, followed by comparison of 31 wild-type respiratory isolates, and showed that employment of sequence typing using the rnpB gene locus was the most specific and reliable. Therefore, the use of rnpB sequencing for the identification of VGS to species level is a reliable and feasible option, based on a single gene target.

Clustered regulatory interspaced short palindromic repeats (CRISPRs) have been discovered in many bacteria and archaea. Many CRISPR-like sequences have been identified in an increasing number of studies on the function of CRISPRs. One CRISPR-like sequence of approximately 240 base pairs has been found to be highly conserved within 11 genome sequences of Streptococcus pneumoniae. A specific CRISPR-like polymerase chain reaction (PCR) assay was designed with the novel primers CRISPR 5F (forward primer) 5'-CTA ATY TCA TAA CCA TAR GAA TC-3' and CRISPR 3R (reverse primer) 5'-GAT AAR ATC CTY TAA WCT TCT AG-3' to detect the presence of this CRISPR-like sequence in pneumococci, as well as in viridans-group streptococci (VGS). This study investigates the prevalence of this CRISPR-like sequence in S. pneumoniae and 12 viridans-group streptococcal species and shows its existence to be shared by the majority of S. pneumoniae and, to a lesser extent, S. mitis. This CRISPR-like sequence was also found in S. australis and it is highly conserved among these strains, suggesting possible biological functional differences from true CRISPR because this CRISPR-like sequence has relatively few repeat numbers, and adjacent homology of CRISPR-associated (cas) genes was absent. The sharing of this CRISPR-like sequence between pneumococci, the mitis group and other VGS, as well as its high sequence homology, may suggest close evolutionary emergence of this sequence between these species.

Forty-eight isolates of viridans-group streptococci (VGS) from adults and children in the community are examined for their resistance to ciprofloxacin phenotypically by determination of the minimum inhibitory concentration (MIC). In addition, the parC gene locus is amplified and sequenced in all isolates and mutations noted. Overall, 44 VGS organisms were found to be susceptible to ciprofloxacin by the broth microdilution method, and the remaining four strains had intermediate susceptibility. Reduced MICs were observed with intermediate strains when reserpine was added to the broth, inhibiting any efflux activity. Overall, the effect of adding reserpine to the broth medium was to add one doubling dilution to the MIC in the case of Streptococcus mitis, S. oralis and S. salivarius, as well as to increase the MIC by two doubling dilutions in two of the three S. parasanguinis isolates. Amino acid sequence analysis of the quinolone resistance-determining region (QRDR) of the parC gene locus showed good correlation to the phenotypic resistance to ciprofloxacin, where no confirmed mutation conferring quinolone resistance was found. Eleven amino acid positions showed discordance with S. pneumoniae R6 and eight (S52, F55, S58, N91, E135, K137, F141 and S167) were common in the VGS species examined. In addition, minor substitutions were found at three positions (D51, T54 and V86). In conclusion, this study demonstrates the low occurrence of ciprofloxacin resistance in a population of VGS isolated from the community. In addition, several silent mutations were noted in VGS organisms without any increase in MIC against ciprofloxacin.

A small study was carried out in order to examine the molecular presence of bla CTX-M gene phylogenetic groups in E. coli (n=263) isolated from food (n=54), water (n=7), animal sources (n=69), using consensus bla CTX-M primers and PCR, in addition to human faecal isolates (n=69) and VTEC O157:H7 (n=64). None of the clinically significant faecal VTEC O157:H7 isolates were shown to carry blaCTX-M type phylogenetic groups, nor were such phylogenetic groups observed in any of the food, water and animal isolates. One community faecal isolate (1/69; 1.4%), dating from 1997, carried this phylogenetic group. As recent work has indicated that a significant proportion of such phylogenetic groups are carried in community isolates of E. coli with little or no hospital contact, it is important that surveillance is increased to identify potential source(s) and reservoirs of such resistance in the community. Further prospective surveillance is thus required to help elucidate the origins of such phylogenetic group in the community. The significance of this study is that the ESBL-producing E. coli associated with local hospital outbreaks is not commonly found in local food, water or animal sources. In addition, given that ESBL-producing E. coli is now a significant organism, both in hospitals and nursing homes in Northern Ireland, this report demonstrates that such organisms were present in the community, as early as 1997.

Polymerase chain reaction amplification of the universal 16S ribosomal RNA (rRNA) gene was performed on a collection of 38 bacterial isolates, originating from air sampled immediately adjacent to the agricultural spreading of bovine slurry. A total of 16 bacterial genera were identified including both Gram-positive and Gram-negative genera. Gram-positive organisms accounted for 34/38 (89.5%) of total bacterial numbers consisting of 12 genera and included Staphylococcus (most common genus isolated), Arthrobacter (2nd most common genus isolated), Brachybacterium, Exiguobacterium, Lactococcus, Microbacterium and Sporosarcina (next most common genera isolated) and finally, Bacillus, Brevibacterium, Frigoribacterium, Mycoplana and Pseudoclavibacter. Gram-negative organisms accounted for only 4/38 (10.5%) bacterial isolates and included the following genera, Brevundimonas, Lysobacter, Psychrobacter and Rhizobium. No gastrointestinal pathogens were detected. Although this study demonstrated a high diversity of the microorganisms present, only a few have been shown to be opportunistically pathogenic to humans and none of these organisms described have been described previously as having an inhalational route of infection and therefore we do not believe that the species of organisms identified pose a significant health and safety threat for immunocompetant individuals., ((c) 2009 Elsevier Inc. All rights reserved.)

Methods employed by the World Health Organization (WHO) are used during this study to determine the optimum storage conditions for maintaining the culturability of Streptococcus pneumoniae in skimmed milk, tryptone, glucose and glycerin (STGG) transport medium. A comparison of S. pneumoniae strains sensitive and resistant to penicillin showed no significant difference in their survival ability in STGG medium. Furthermore, it is confirmed that storage at -70 degrees C remains most effective for maintaining viability by culture of S. pneumoniae. Storage at -20 degrees C would only be acceptable in the short-term, while storage at +4 degrees C is not recommended. Of note, this study has shown STGG medium at room temperature to be an efficient growth medium for pneumococci in the short-term. This work will help to establish robust sampling protocols when performing community studies to ensure culturability of comparison between community and laboratory pneumococci survival.

MicroRNAs are a class of small non-coding RNAs widely reported in eukaryotic multicellular organisms. In this study, a number of small non-coding micro (mi)RNA species in clinical isolates of prokaryote Pseudomonas aeruginosa were obtained from the sputum of adult patients with cystic fibrosis (CF) utilising a DynaExpress miRNA cloning kit, and five miRNAs of 16-47 nucleotides that were smaller than those encountered or described (80-100 nucleotides) previously in bacterial systems were described. This report presents data on these unknown cellular miRNAs cloned from P. aeruginosa isolates from CF patients. Adapting a computational miRNA prediction model that takes advantage of the highly conserved known miRNA hair pin stems regions, the results revealed that the fold structure of the microRNAs had a high homology to the recently reported human bacterial infection response (BiR)-related microRNA, mi-146, associated with the Toll-like receptor (TLR) family, which is the primary evolutionarily conserved sensors of pathogen-associated molecular patterns (PAMPs), and known to trigger host inflammatory and immune responses.

Molecular epidemiology of verocytoxigenic Escherichia coli O157:H7 is important to help elucidate reservoirs and modes of transmission, particularly between animals and humans. As the recA gene locus is now beginning to gain application in bacterial genotyping schemes, and as it has not been examined previously in E. coli O157 isolates, this study aims to examine potential polymorphic variation as a possible epidemiological marker for the subspecies characterisation of clinically significant verocytotoxigenic E. coli O157:H7. A novel polymerase chain reaction (PCR) assay was designed to target a 638 bp region of the recA gene in E. coli O157 isolates. The PCR amplification of genomic DNA from extracted organisms was able to generate an amplicon of the expected size (approximately 638 bp) for all E. coli O157:H7 examined (n=80), as well as for other non-O157 E. coli and other members of the Enterobacteriaeceae including Citrobacter, Hafnia, Shigella, Enterobacter and Providencia. Subsequent restriction fragment length polymorphism (RFLP) and single-stranded conformational polymorphism (SSCP) analyses of these recA amplicons were able to differentiate E. coli O157 from the organisms examined, but were unable to distinguish between 79 isolates of wild-type E. coli O157, suggesting a highly conserved recA gene structure within the local population of organisms examined.

Traditional herbal cures and remedies have played an important historical role in the treatment of a variety of illnesses and diseases in Northern Ireland for the last three hundred years. Recently, these have been reviewed in the publication by Linda Ballard from the Ulster Folk and Transport Museum at Cultra, Co. Down, which details the variety of local plants used and for what purpose. From this publication and another related publication, we note the description of several plant species that consistently appear in traditional cures and remedies, particularly used to treat infections and infectious diseases. Unfortunately, although these plants have strong associations with the local historical evidence base, there are very limited and mainly no formal publications in the medical/scientific evidence base, examining their scientific background and clinical efficacy.