Cystic Fibrosis-Related Gut Dysbiosis: A Systematic Review.

BACKGROUND AND AIMS: Cystic Fibrosis (CF) is associated with gut dysbiosis, local and systemic inflammation, and impaired immune function. Gut microbiota dysbiosis results from changes in the complex gut milieu in response to CF transmembrane conductance regulator (CFTR) dysfunction, pancreatic malabsorption, diet, medications, and environmental influences. In several diseases, alteration of the gut microbiota influences local and systemic inflammation and disease outcomes. We conducted a systematic review of the gut microbiota in CF and explored factors influencing dysbiosis. METHODS: An electronic search of three databases was conducted in January 2019, and re-run in June 2021. Human, animal, and in vitro studies were included. The primary outcome was differences in the gut microbiota between people with CF (pwCF) and healthy controls. Secondary outcomes included the relationship between the gut microbiota and other factors, including diet, medication, inflammation, and pulmonary function in pwCF. RESULTS: Thirty-eight studies were identified. The literature confirmed the presence of CF-related gut dysbiosis, characterized by reduced diversity and several taxonomic changes. There was a relative increase of bacteria associated with a pro-inflammatory response coupled with a reduction of those considered anti-inflammatory. However, studies linking gut dysbiosis to systemic and lung inflammation were limited. Causes of gut dysbiosis were multifactorial, and findings were variable. Data on the impact of CFTR modulators on the gut microbiota were limited. CONCLUSIONS: CF-related gut dysbiosis is evident in pwCF. Whether this influences local and systemic disease and is amenable to interventions with diet and drugs, such as CFTR modulators, requires further investigation.

Staphylococcus pseudoxylosus sp. nov., isolated from bovine mastitis.

Strain S04009T, a Gram-stain-positive, coagulase-negative staphylococcus, was isolated from bovine mastitis in France. 16S rRNA gene analysis revealed it to be closely related to the coagulase-negative species Staphylococcusxylosus, Staphylococcussaprophyticus, Staphylococcuscaeli and Staphylococcus edaphicus. At the whole-genome level, strain S04009T had an average nucleotide identity value <95 % and an inferred DNA-DNA hybridization value <70 % when compared to these species. Furthermore, phenotypic characteristics distinguished S04009T from those species. From these related species only strain S04009T and S. xylosus are able to ferment xylose and these two can be distinguished by the inability of strain S04009T to express urease activity. Based on the genotypic and phenotypic results, it is proposed that this isolate is a novel species, with the name Staphylococcus pseudoxylosus sp. nov. The type strain is S04009T (=DSM 107950T=CCUG 72763T=NCTC 14184T).

Staphylococcus caeli sp. nov., isolated from air sampling in an industrial rabbit holding.

Strain 82T, a Gram-stain-positive, coagulase-negative staphylococcus was isolated from an air sample obtained from an industrial rabbit holding in Italy. It is phylogenetically closely related to the coagulase-negative species Staphylococcus saprophyticus, Staphylococcus xylosus and Staphylococcus edaphicus. However, it could be distinguished from these species by sequence differences between the 16S rRNA, hsp60, rpoB, dnaJ and gap genes. At the whole genome level, the isolate had an average nucleotide identity of <95 % and an inferred DNA-DNA hybridization of <70 % when compared to these species. Based on the genotypic results, it is proposed that this isolate is a novel species, with the name Staphylococcus caeli sp. nov. The type strain is 82BT (=NCTC 14063T=CCUG 71912T).

A mecC allotype, mecC3, in the CoNS Staphylococcus caeli, encoded within a variant SCCmecC.

BACKGROUND: Methicillin resistance in staphylococci is conferred by an alternative PBP (PBP2a/2') with low affinity for most ß-lactam antibiotics. PBP2a is encoded by mecA, which is carried on a mobile genetic element known as SCCmec. A variant of mecA, mecC, was described in 2011 and has been found in Staphylococcus aureus from humans and a wide range of animal species as well as a small number of other staphylococcal species from animals. OBJECTIVES: We characterized a novel mecC allotype, mecC3, encoded by an environmental isolate of Staphylococcus caeli cultured from air sampling of a commercial rabbit holding. METHODS: The S. caeli isolate 82BT was collected in Italy in 2013 and genome sequenced using MiSeq technology. This allowed the assembly and comparative genomic study of the novel SCCmec region encoding mecC3. RESULTS: The study isolate encodes a novel mecA allotype, mecC3, with 92% nucleotide identity to mecC. mecC3 is encoded within a novel SCCmec element distinct from those previously associated with mecC, including a ccrAB pairing (ccrA5B3) not previously linked to mecC. CONCLUSIONS: This is the first description of the novel mecC allotype mecC3, the first isolation of a mecC-positive Staphylococcus in Italy and the first report of mecC in S. caeli. Furthermore, the SCCmec element described here is highly dissimilar to the archetypal SCCmec XI encoding mecC in S. aureus and to elements encoding mecC in other staphylococci. Our report highlights the diversity of mecC allotypes and the diverse staphylococcal species, ecological settings and genomic context in which mecC may be found.

The widespread use of topical antimicrobials enriches for resistance in Staphylococcus aureus isolated from patients with atopic dermatitis.

BACKGROUND: Carriage rates of Staphylococcus aureus on affected skin in atopic dermatitis (AD) are approximately 70%. Increasing disease severity during flares and overall disease severity correlate with increased burden of S. aureus. Treatment in AD therefore often targets S. aureus with topical and systemic antimicrobials. OBJECTIVES: To determine whether antimicrobial sensitivities and genetic determinants of resistance differed in S. aureus isolates from the skin of children with AD and healthy child nasal carriers. METHODS: In this case-control study, we compared S. aureus isolates from children with AD (n = 50) attending a hospital dermatology department against nasal carriage isolates from children without skin disease (n = 49) attending a hospital emergency department for noninfective conditions. Using whole genome sequencing we generated a phylogenetic framework for the isolates based on variation in the core genome, then compared antimicrobial resistance phenotypes and genotypes between disease groups. RESULTS: Staphylococcus aureus from cases and controls had on average similar numbers of phenotypic resistances per isolate. Case isolates differed in their resistance patterns, with fusidic acid resistance (FusR ) being significantly more frequent in AD (P = 0·009). The genetic basis of FusR also differentiated the populations, with chromosomal mutations in fusA predominating in AD (P = 0·049). Analysis revealed that FusR evolved multiple times and via multiple mechanism in the population. Carriage of plasmid-derived qac genes, which have been associated with reduced susceptibility to antiseptics, was eight times more frequent in AD (P = 0·016). CONCLUSIONS: The results suggest that strong selective pressure drives the emergence and maintenance of specific resistances in AD.

Low genomic diversity of Legionella pneumophila within clinical specimens.

OBJECTIVES: Legionella pneumophila is the leading cause of Legionnaires' disease, a severe form of pneumonia acquired from environmental sources. Investigations of both sporadic cases and outbreaks rely mostly on analysis of a single to a few colony pick(s) isolated from each patient. However, because of the lack of data describing diversity within single patients, the optimal number of picks is unknown. Here, we investigated diversity within individual patients using sequence-based typing (SBT) and whole-genome sequencing (WGS). METHODS: Ten isolates of L. pneumophila were obtained from each of ten epidemiologically unrelated patients. SBT and WGS were undertaken, and single-nucleotide polymorphisms (SNPs) were identified between isolates from the same patient. RESULTS: The same sequence type (ST) was obtained for each set of ten isolates. Using genomic analysis, zero SNPs were identified between isolates from seven patients, a maximum of one SNP was found between isolates from two patients, and a maximum of two SNPs was found amongst isolates from one patient. Assuming that the full within-host diversity has been captured with ten isolates, statistical analyses showed that, on average, analysis of one isolate would yield a 70% chance of capturing all observed genotypes, and seven isolates would yield a 90% chance. CONCLUSIONS: SBT and WGS analyses of multiple colony picks obtained from ten patients showed no, or very low, within-host genomic diversity in L. pneumophila, suggesting that analysis of one colony pick per patient will often be sufficient to obtain reliable typing data to aid investigation of cases of Legionnaires' disease.

Rapid detection and evolutionary analysis of Legionella pneumophila serogroup 1 sequence type 47.

OBJECTIVES: Legionella pneumophila serogroup 1 (Lp1) sequence type 47 is the leading cause of legionellosis in north-western Europe, but, surprisingly, it is rarely isolated from environmental samples. Comparative genomics was applied to develop a PCR assay and to better understand the evolution of this strain. METHODS: Comparative analysis of 36 genomes representative of the Lp species was used to identify specific PCR targets, which were then evaluated in silico on 545 sequenced genomes and in vitro on 436 Legionella strains, 106 respiratory samples, and three environmental samples from proven ST47 sources. Phylogenetic analyses were performed to understand the evolution of ST47. RESULTS: The gene LPO_1073 was characterized as being 100% conserved in all 129 ST47 genomes analysed. A real-time PCR designed to detect LPO_1073 was positive for all 110 ST47 strains tested and agreed with culture and typing results previously obtained for 106 respiratory samples. The three environmental samples were also positive. Surprisingly, 26 of the 44 ST109 strains tested among 342 non-ST47 strains scored positive for LPO_1073. SNP-based phylogenetic analysis was undertaken to understand this result: the PCR-positive ST109 genomes were almost identical to ST47 genomes, with the exception of a recombined region probably acquired by ST47 from a ST62(-like) strain. CONCLUSION: The genomic analysis allowed the design of a highly specific PCR assay for rapid detection of ST47 strains. Furthermore, it allowed us to uncover the evolution of ST47 strains from ST109 by homologous recombination with ST62. We hypothesize that this recombination generated the leading cause of legionellosis in north-western Europe.

Community dynamics and the lower airway microbiota in stable chronic obstructive pulmonary disease, smokers and healthy non-smokers.

RATIONALE: The role bacteria play in the progression of COPD has increasingly been highlighted in recent years. However, the microbial community complexity in the lower airways of patients with COPD is poorly characterised. OBJECTIVES: To compare the lower airway microbiota in patients with COPD, smokers and non-smokers. METHODS: Bronchial wash samples from adults with COPD (n=18), smokers with no airways disease (n=8) and healthy individuals (n=11) were analysed by extended-culture and culture-independent Illumina MiSeq sequencing. We determined aerobic and anaerobic microbiota load and evaluated differences in bacteria associated with the three cohorts. Culture-independent analysis was used to determine differences in microbiota between comparison groups including taxonomic richness, diversity, relative abundance, 'core' microbiota and co-occurrence. MEASUREMENT AND MAIN RESULTS: Extended-culture showed no difference in total load of aerobic and anaerobic bacteria between the three cohorts. Culture-independent analysis revealed that the prevalence of members of Pseudomonas spp. was greater in the lower airways of patients with COPD; however, the majority of the sequence reads for this taxa were attributed to three patients. Furthermore, members of Bacteroidetes, such as Prevotella spp., were observed to be greater in the 'healthy' comparison groups. Community diversity (α and ß) was significantly less in COPD compared with healthy groups. Co-occurrence of bacterial taxa and the observation of a putative 'core' community within the lower airways were also observed. CONCLUSIONS: Microbial community composition in the lower airways of patients with COPD is significantly different to that found in smokers and non-smokers, indicating that a component of the disease is associated with changes in microbiological status.

Variable alterations of the microbiota, without metabolic or immunological change, following faecal microbiota transplantation in patients with chronic pouchitis.

Faecal microbiota transplantation (FMT) is effective in the treatment of Clostridium difficile infection, where efficacy correlates with changes in microbiota diversity and composition. The effects of FMT on recipient microbiota in inflammatory bowel diseases (IBD) remain unclear. We assessed the effects of FMT on microbiota composition and function, mucosal immune response, and clinical outcome in patients with chronic pouchitis. Eight patients with chronic pouchitis (current PDAI ≥7) were treated with FMT via nasogastric administration. Clinical activity was assessed before and four weeks following FMT. Faecal coliform antibiotic sensitivities were analysed, and changes in pouch faecal and mucosal microbiota assessed by 16S rRNA gene pyrosequencing and (1)H NMR spectroscopy. Lamina propria dendritic cell phenotype and cytokine profiles were assessed by flow cytometric analysis and multiplex assay. Following FMT, there were variable shifts in faecal and mucosal microbiota composition and, in some patients, changes in proportional abundance of species suggestive of a "healthier" pouch microbiota. However, there were no significant FMT-induced metabolic or immunological changes, or beneficial clinical response. Given the lack of clinical response following FMT via a single nasogastric administration our results suggest that FMT/bacteriotherapy for pouchitis patients requires further optimisation.

Detection of livestock-associated meticillin-resistant Staphylococcus aureus CC398 in retail pork, United Kingdom, February 2015.

Livestock-associated meticillin-resistant Staphylococcus aureus belonging to clonal complex 398 (LA-MRSA CC398) is an important cause of zoonotic infections in many countries. Here, we describe the isolation of LA-MRSA CC398 from retail meat samples of United Kingdom (UK) farm origin. Our findings indicate that this lineage is probably established in UK pig farms and demonstrate a potential pathway for the transmission of LA-MRSA CC398 from livestock to humans in the UK.

Large-scale whole genome sequencing of M. tuberculosis provides insights into transmission in a high prevalence area.

To improve understanding of the factors influencing tuberculosis transmission and the role of pathogen variation, we sequenced all available specimens from patients diagnosed over 15 years in a whole district in Malawi. Mycobacterium tuberculosis lineages were assigned and transmission networks constructed, allowing ≤10 single nucleotide polymorphisms (SNPs) difference. We defined disease as due to recent infection if the network-determined source was within 5 years, and assessed transmissibility from forward transmissions resulting in disease. High-quality sequences were available for 1687 disease episodes (72% of all culture-positive episodes): 66% of patients linked to at least one other patient. The between-patient mutation rate was 0.26 SNPs/year (95% CI 0.21-0.31). We showed striking differences by lineage in the proportion of disease due to recent transmission and in transmissibility (highest for lineage-2 and lowest for lineage-1) that were not confounded by immigration, HIV status or drug resistance. Transmissions resulting in disease decreased markedly over time.

Prevalence and characterization of human mecC methicillin-resistant Staphylococcus aureus isolates in England.

OBJECTIVES: There are limited data available on the epidemiology and prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in the human population that encode the recently described mecA homologue, mecC. To address this knowledge gap we undertook a prospective prevalence study in England to determine the prevalence of mecC among MRSA isolates. PATIENTS AND METHODS: Three hundred and thirty-five sequential MRSA isolates from individual patients were collected from each of six clinical microbiology laboratories in England during 2011-12. These were tested by PCR or genome sequencing to differentiate those encoding mecA and mecC. mecC-positive isolates were further characterized by multilocus sequence typing, spa typing, antimicrobial susceptibility profile and detection of PBP2a using commercially available kits. RESULTS: Nine out of the 2010 MRSA isolates tested were mecC positive, indicating a prevalence among MRSA in England of 0.45% (95% CI 0.24%-0.85%). The remainder were mecA positive. Eight out of these nine mecC MRSA isolates belonged to clonal complex 130, the other being sequence type 425. Resistance to non-ß-lactam antibiotics was rare among these mecC MRSA isolates and all were phenotypically identified as MRSA using oxacillin and cefoxitin according to BSAC disc diffusion methodology. However, all nine mecC isolates gave a negative result using three different commercial PBP2a detection assays. CONCLUSIONS: mecC MRSA are currently rare among MRSA isolated from humans in England and this study provides an important baseline prevalence rate to monitor future changes, which may be important given the increasing prevalence of mecC MRSA reported in Denmark.

Prevalence and properties of mecC methicillin-resistant Staphylococcus aureus (MRSA) in bovine bulk tank milk in Great Britain.

OBJECTIVES: mecC methicillin-resistant Staphylococcus aureus (MRSA) represent a newly recognized form of MRSA, distinguished by the possession of a divergent mecA homologue, mecC. The first isolate to be identified came from bovine milk, but there are few data on the prevalence of mecC MRSA among dairy cattle. The aim of this study was to conduct a prevalence study of mecC MRSA among dairy farms in Great Britain. METHODS: Test farms were randomly selected by random order generation and bulk tank samples were tested for the presence of mecC MRSA by broth enrichment and plating onto chromogenic agar. All MRSA isolated were screened by PCR for mecA and mecC, and mecC MRSA were further characterized by multilocus sequence typing, spa typing and antimicrobial susceptibility testing. RESULTS: mecC MRSA were detected on 10 of 465 dairy farms sampled in England and Wales (prevalence 2.15%, 95% CI 1.17%-3.91%), but not from 625 farms sampled in Scotland (95% CI of prevalence 0%-0.61%). Seven isolates belonged to sequence type (ST) 425, while the other three belonged to clonal complex 130. Resistance to non-ß-lactam antibiotics was uncommon. All 10 isolates produced a negative result by slide agglutination for penicillin-binding protein 2a. mecA MRSA ST398 was detected on one farm in England. CONCLUSIONS: mecC MRSA is widely distributed among dairy farms in Great Britain, but this distribution is not uniform across the whole country. These results provide an important baseline dataset to monitor the epidemiology of this emerging form of MRSA.

Distinguishable epidemics of multidrug-resistant Salmonella Typhimurium DT104 in different hosts.

The global epidemic of multidrug-resistant Salmonella Typhimurium DT104 provides an important example, both in terms of the agent and its resistance, of a widely disseminated zoonotic pathogen. Here, with an unprecedented national collection of isolates collected contemporaneously from humans and animals and including a sample of internationally derived isolates, we have used whole-genome sequencing to dissect the phylogenetic associations of the bacterium and its antimicrobial resistance genes through the course of an epidemic. Contrary to current tenets supporting a single homogeneous epidemic, we demonstrate that the bacterium and its resistance genes were largely maintained within animal and human populations separately and that there was limited transmission, in either direction. We also show considerable variation in the resistance profiles, in contrast to the largely stable bacterial core genome, which emphasizes the critical importance of integrated genotypic data sets in understanding the ecology of bacterial zoonoses and antimicrobial resistance.

Unusual features in organisation of capsular polysaccharide-related genes of C. jejuni strain X.

PCR probing of the genome of Campylobacter jejuni strain X using conserved capsular polysaccharide (CPS)-related genes allowed elucidation of a complete sequence of the respective gene cluster (cps). This is the largest known Campylobacter cps cluster (38 kb excluding flanking kps regions), which includes a number of genes not detected in other Campylobacter strains. Sequence analysis suggests genetic rearrangements both within and outside the cps gene cluster, a mechanism which may be responsible for mosaic organisation of sugar transferase-related genes leading to structural variability of the capsular polysaccharide (CPS).

First detection of livestock-associated meticillin-resistant Staphylococcus aureus CC398 in bulk tank milk in the United Kingdom, January to July 2012.

Livestock-associated meticillin-resistant Staphylococcus aureus belonging to clonal complex 398 (LA-MRSA CC398) is an important cause of zoonotic infections in several countries, but there is only a single published report of this lineage from the United Kingdom (UK). Here, we describe the isolation of LA-MRSA CC398 from bulk tank milk from five geographically dispersed farms in the UK. Our findings suggest that LA-MRSA CC398 is established in livestock in the UK. Awareness of the potential occupational risks and surveillance in other food-producing animal species should be promoted.

The newly described mecA homologue, mecALGA251, is present in methicillin-resistant Staphylococcus aureus isolates from a diverse range of host species.

OBJECTIVES: A previously unidentified mecA homologue, mecA(LGA251), has recently been described in methicillin-resistant Staphylococcus aureus (MRSA) from humans and dairy cattle. The origin and epidemiology of this novel homologue are unclear. The objective of this study was to provide basic descriptive information of MRSA isolates harbouring mecA(LGA251) from a range of host animal species. METHODS: A number of S. aureus isolates from historical animal isolate collections were chosen for investigation based on their similarity to known mecA(LGA251) MRSA isolates. The presence of mecA(LGA251) was determined using a multiplex PCR and antimicrobial susceptibility testing performed by disc diffusion. RESULTS: MRSA harbouring mecA(LGA251) were found in isolates from a domestic dog, brown rats, a rabbit, a common seal, sheep and a chaffinch. All of the isolates were phenotypically MRSA, although this depended on which test was used; some isolates would be considered susceptible with certain assays. All isolates were susceptible to linezolid, rifampicin, kanamycin, norfloxacin, erythromycin, clindamycin, fusidic acid, tetracycline, trimethoprim/sulfamethoxazole and mupirocin. Five multilocus sequence types were represented (2273, 130, 425, 1764 and 1245) and six spa types (t208, t6293, t742, t6594, t7914 and t843). CONCLUSIONS: The discovery of MRSA isolates possessing mecA(LGA251) from a diverse range of host species, including different taxonomic classes, has important implications for the diagnosis of MRSA in these species and our understanding of the epidemiology of this novel mecA homologue.

Complete genome sequence of the plant pathogen Erwinia amylovora strain ATCC 49946.

Erwinia amylovora causes the economically important disease fire blight that affects rosaceous plants, especially pear and apple. Here we report the complete genome sequence and annotation of strain ATCC 49946. The analysis of the sequence and its comparison with sequenced genomes of closely related enterobacteria revealed signs of pathoadaptation to rosaceous hosts.