Community methicillin-resistant Staphylococcus aureus outbreaks in areas of low prevalence.

BACKGROUND: Community-acquired (CA), community-onset methicillin-resistant Staphylococcus aureus (CO-MRSA) infection presents a significant public health challenge, even where MRSA rates are historically lower. Despite successes in reducing hospital-onset MRSA, CO-MRSA rates are increasing globally, with a need to understand this trend, and the potential risk factors for re-emergence. OBJECTIVES: This review aims to explore the characteristics of outbreaks of community-acquired community-onset methicillin-resistant Staphylococcus aureus in low-prevalence areas, to understand the factors involved in its rise, and to translate this knowledge into public health policy and further research needs. SOURCES: PubMed, EMBASE, and Google Scholar were searched using combinations of the terms 'transmission', 'acquisition', 'community-acquired', 'MRSA', 'CA-MRSA', 'low prevalence', 'genomic', 'outbreak', 'colonisation', and 'carriage'. Wherever evidence was limited, additional articles were sought specifically, via PubMed searches. Papers where materials were not available in English were excluded. CONTENT: Challenges in defining low-prevalence areas and the significance of exposure to various risk factors for community acquisition, such as healthcare settings, travel, livestock, and environmental factors, are discussed. The importance of genomic surveillance in identifying outbreak strains and understanding the transmission dynamics is highlighted, along with the need for robust public health policies and control measures. IMPLICATIONS: The findings emphasise the complexity of CO-MRSA transmission and the necessity of a multifaceted approach in low-prevalence areas. This includes integrated and systematic surveillance of hospital-onset-, CO-, and livestock-associated MRSA, as has been effective in some Northern European countries. The evolution of CO-MRSA underscores the need for global collaboration, routine genomic surveillance, and comprehensive antimicrobial stewardship to mitigate the rise of CO-MRSA and address the broader challenge of antimicrobial resistance. These efforts are crucial for maintaining low MRSA prevalence and managing the increasing burden of CO-MRSA in both low and higher prevalence regions.

Optimization of high-throughput 16S rRNA gene amplicon sequencing: an assessment of PCR pooling, mastermix use and contamination.

16S rRNA gene sequencing is widely used to characterize human and environmental microbiomes. Sequencing at scale facilitates better powered studies but is limited by cost and time. We identified two areas in our 16S rRNA gene library preparation protocol where modifications could provide efficiency gains, including (1) pooling of multiple PCR amplifications per sample to reduce PCR drift and (2) manual preparation of mastermix to reduce liquid handling. Using nasal samples from healthy human participants and a serially diluted mock microbial community, we compared alpha and beta diversity, and compositional abundance where the PCR amplification was conducted in triplicate, duplicate or as a single reaction, and where manually prepared or premixed mastermix was used. One hundred and fifty-eight 16S rRNA gene sequencing libraries were prepared, including a replicate experiment. Comparing PCR pooling strategies, we found no significant difference in high-quality read counts and alpha diversity, and beta diversity by Bray-Curtis index clustered by replicate on principal coordinate analysis (PCoA) and non-metric dimensional scaling (NMDS) analysis. Choice of mastermix had no significant impact on high-quality read and alpha diversity, and beta diversity by Bray-Curtis index clustered by replicate in PCoA and NMDS analysis. Importantly, we observed contamination and variability of rare species (<0.01 %) across replicate experiments; the majority of contaminants were accounted for by removal of species present at <0.1 %, or were linked to reagents (including a primer stock). We demonstrate no requirement for pooling of PCR amplifications or manual preparation of PCR mastermix, resulting in a more efficient 16S rRNA gene PCR protocol.

Genomic identification of cryptic susceptibility to penicillins and ß-lactamase inhibitors in methicillin-resistant Staphylococcus aureus.

Antibiotic resistance in bacterial pathogens threatens the future of modern medicine. One such resistant pathogen is methicillin-resistant Staphylococcus aureus (MRSA), which is resistant to nearly all ß-lactam antibiotics, limiting treatment options. Here, we show that a significant proportion of MRSA isolates from different lineages, including the epidemic USA300 lineage, are susceptible to penicillins when used in combination with ß-lactamase inhibitors such as clavulanic acid. Susceptibility is mediated by a combination of two different mutations in the mecA promoter region that lowers mecA-encoded penicillin-binding protein 2a (PBP2a) expression, and in the majority of isolates by either one of two substitutions in PBP2a (E246G or M122I) that increase the affinity of PBP2a for penicillin in the presence of clavulanic acid. Treatment of S. aureus infections in wax moth and mouse models shows that penicillin/ß-lactamase inhibitor susceptibility can be exploited as an effective therapeutic choice for 'susceptible' MRSA infection. Finally, we show that isolates with the PBP2a E246G substitution have a growth advantage in the presence of penicillin but the absence of clavulanic acid, which suggests that penicillin/ß-lactamase susceptibility is an example of collateral sensitivity (resistance to one antibiotic increases sensitivity to another). Our findings suggest that widely available and currently disregarded antibiotics could be effective in a significant proportion of MRSA infections.