Cutibacterium avidum: A Potent and Underestimated Pathogen in Prosthetic Hip Joint Infections.

Cutibacterium avidum has recently been reported as a rare cause of prosthetic joint infections (PJIs), contrary to Cutibacterium acnes, which is well established as a cause of PJIs, especially in shoulder arthroplasties. Two specific risk factors for PJI due to C. avidum have been reported: obesity and the skin incision approach. Here, we report four cases of hip PJIs caused by C. avidum admitted over a 30-month period at a single center. Whole-genome sequencing revealed that the four C. avidum strains were all individual strains and did not originate from a common source, such as an outbreak. Antibiotic susceptibility tests showed that the isolates were fully susceptible, and none carried known antibiotic resistance genes. In conclusion, the occurrence of four cases of PJI caused by C. avidum over a limited time at a single center may indicate that this pathogen is underestimated and is either emerging or more common than previously recognized. The patients presented overt signs of infection during surgery, indicating that C. avidum is a virulent pathogen. None of the previously reported risk factors for C. avidum PJI applied to these patients as only one was obese and none were operated on using a direct anterior skin incision approach.

Delivery of a sebum modulator by an engineered skin microbe in mice.

Microorganisms can be equipped with synthetic genetic programs for the production of targeted therapeutic molecules. Cutibacterium acnes is the most abundant commensal of the human skin, making it an attractive chassis to create skin-delivered therapeutics. Here, we report the engineering of this bacterium to produce and secrete the therapeutic molecule neutrophil gelatinase-associated lipocalin, in vivo, for the modulation of cutaneous sebum production.

Species- and strain-level diversity of Corynebacteria isolated from human facial skin.

BACKGROUND: Sequencing of the human skin microbiome revealed that Corynebacterium is an ubiquitous and abundant bacterial genus on human skin. Shotgun sequencing further highlighted the microbial "dark matter" of the skin microbiome, consisting of microorganisms, including corynebacterial species that were not cultivated and genome-sequenced so far. In this pilot project, facial human skin swabs of 13 persons were cultivated to selectively obtain corynebacteria. 54 isolates were collected and 15 of these were genome-sequenced and the pan-genome was determined. The strains were biochemically characterized and antibiotic susceptibility testing (AST) was performed. RESULTS: Among the 15 sequenced strains, nine different corynebacterial species were found, including two so far undescribed species, tentatively named "Corynebacterium vikingii" and "Corynebacterium borealis", for which closed genome sequences were obtained. Strain variability beyond the species level was determined in biochemical tests, such as the variable presence of urease activity and the capacity to ferment different sugars. The ability to grow under anaerobic conditions on solid agar was found to be species-specific. AST revealed resistances to clindamycin in seven strains. A Corynebacterium pseudokroppenstedtii strain showed additional resistance towards beta-lactam and fluoroquinolone antibiotics; a chromosomally located 17 kb gene cluster with five antibiotic resistance genes was found in the closed genome of this strain. CONCLUSIONS: Taken together, this pilot study identified an astonishing diversity of cutaneous corynebacterial species in a relatively small cohort and determined species- and strain-specific individualities regarding biochemical and resistance profiles. This further emphasizes the need for cultivation-based studies to be able to study these microorganisms in more detail, in particular regarding their host-interacting and, potentially, -beneficial and/or -detrimental properties.

The value of sonication on orthopaedic implants in an everyday clinical setting - an exploratory study.

BACKGROUND: Sonication of removed orthopaedic implants in suspected implant-associated infections (IAI) is widely applied internationally. However, evaluation of the utility of sonication on all implants removed in everyday standard practice is scarce. This exploratory study was performed to evaluate the application of sonication fluid (SF) culture on removed orthopaedic implants, irrespective of the reason for removal. METHODS: Out of 100 removed orthopaedic implants collected between August 2019 and September 2020, 77 implants with availability of concurrent tissue culture samples were included in the study. Removed implants were categorized into a confirmed or suspected IAI group and a presumed aseptic group based on pre-operative diagnosis by the responsible surgeon. Implants were sonicated and SF culture performed under both aerobic and anaerobic conditions. The significance of all bacterial isolates was evaluated based on the CFU/mL cut-offs of the EBJIS guidelines, except for C. acnes where additional investigations were performed. RESULTS: The results of SF culture in the two groups were compared with their corresponding tissue cultures. Out of the 12 cases in the confirmed/suspected IAI group, SF culture was positive in 11 cases and had increased diagnostic yield in two (17%) cases compared to tissue culture. Increased diagnostic yield of SF compared to tissue culture was seen in seven (11%) of the 65 implants in the presumed aseptic group. If growth of Cutibacterium species isolates were interpreted based on EBJIS cut-off for SF culture instead of the study-specific criteria, then two isolates considered to represent infection might have been missed while three other isolates considered contaminants would have fallen under the 'infection confirmed' category in the EBJIS guidelines. CONCLUSION: Sonication with SF culture has increased diagnostic yield compared to tissue cultures in all implants irrespective of reason for removal. However, positive SF cultures with Cutibacterium species should always be interpreted with extreme care.

Shotgun sequencing of sonication fluid for the diagnosis of orthopaedic implant-associated infections with Cutibacterium acnes as suspected causative agent.

Orthopaedic implant-associated infections (OIAIs) due to Cutibacterium acnes can be difficult to diagnose. The aim of this pilot study was to determine if metagenomic next-generation sequencing (mNGS) can provide additional information to improve the diagnosis of C. acnes OIAIs. mNGS was performed on sonication fluid (SF) specimens derived from 24 implants. These were divided into three groups, based on culture results: group I, culture-negative (n = 4); group II, culture-positive for C. acnes (n = 10); and group III, culture-positive for other bacteria (n = 10). In group I, sequence reads from C. acnes were detected in only one SF sample, originating from a suspected case of OIAIs, which was SF and tissue culture-negative. In group II, C. acnes sequences were detected in 7/10 samples. In group III, C. acnes sequence reads were found in 5/10 samples, in addition to sequence reads that matched the bacterial species identified by culture. These samples could represent polymicrobial infections that were missed by culture. Taken together, mNGS was able to detect C. acnes DNA in more samples compared to culture and could be used to identify cases of suspected C. acnes OIAIs, in particular regarding possible polymicrobial infections, where the growth of C. acnes might be compromised due to a fast-growing bacterial species. However, since SF specimens are usually low-biomass samples, mNGS is prone to DNA contamination, possibly introduced during DNA extraction or sequencing procedures. Thus, it is advisable to set a sequence read count threshold, taking into account project- and NGS-specific criteria.

Biofilm formation and inflammatory potential of Staphylococcus saccharolyticus: A possible cause of orthopedic implant-associated infections.

Staphylococcus saccharolyticus, a coagulase-negative staphylococcal species, has some unusual characteristics for human-associated staphylococci, such as slow growth and its preference for anoxic culture conditions. This species is a relatively abundant member of the human skin microbiota, but its microbiological properties, as well as the pathogenic potential, have scarcely been investigated so far, despite being occasionally isolated from different types of infections including orthopedic implant-associated infections. Here, we investigated the growth and biofilm properties of clinical isolates of S. saccharolyticus and determined host cell responses. Growth assessments in anoxic and oxic conditions revealed strain-dependent outcomes, as some strains can also grow aerobically. All tested strains of S. saccharolyticus were able to form biofilm in a microtiter plate assay. Strain-dependent differences were determined by optical coherence tomography, revealing that medium supplementation with glucose and sodium chloride enhanced biofilm formation. Visualization of the biofilm by confocal laser scanning microscopy revealed the role of extracellular DNA in the biofilm structure. In addition to attached biofilms, S. saccharolyticus also formed bacterial aggregates at an early stage of growth. Transcriptome analysis of biofilm-grown versus planktonic cells revealed a set of upregulated genes in biofilm-embedded cells, including factors involved in adhesion, colonization, and competition such as epidermin, type I toxin-antitoxin system, and phenol-soluble modulins (beta and epsilon). To investigate consequences for the host after encountering S. saccharolyticus, cytokine profiling and host cell viability were assessed by infection experiments with differentiated THP-1 cells. The microorganism strongly triggered the secretion of the tested pro-inflammatory cyto- and chemokines IL-6, IL-8, and TNF-alpha, determined at 24 h post-infection. S. saccharolyticus was less cytotoxic than Staphylococcus aureus. Taken together, the results indicate that S. saccharolyticus has substantial pathogenic potential. Thus, it can be a potential cause of orthopedic implant-associated infections and other types of deep-seated infections.

Interference and co-existence of staphylococci and Cutibacterium acnes within the healthy human skin microbiome.

Human skin is populated by trillions of microbes collectively called the skin microbiome. Staphylococcus epidermidis and Cutibacterium acnes are among the most abundant members of this ecosystem, with described roles in skin health and disease. However, knowledge regarding the health beneficial effects of these ubiquitous skin residents is still limited. Here, we profiled the staphylococcal and C. acnes landscape across four different skin sites of 30 individuals (120 skin samples) using amplicon-based next-generation sequencing. Relative abundance profiles obtained indicated the existence of phylotype-specific co-existence and exclusion scenarios. Co-culture experiments with 557 staphylococcal strains identified 30 strains exhibiting anti-C. acnes activities. Notably, staphylococcal strains were found to selectively exclude acne-associated C. acnes and co-exist with healthy skin-associated phylotypes, through regulation of the antimicrobial activity. Overall, these findings highlight the importance of skin-resident staphylococci and suggest that selective microbial interference is a contributor to healthy skin homeostasis.

Microbiological Characterization of Cutibacterium acnes Strains Isolated from Prosthetic Joint Infections.

Aims: This study aimed to characterize 79 Cutibacterium acnes strains isolated from prosthetic joint infections (PJIs) originated from eight European hospitals. Methods: Isolates were phylotyped according to the single-locus sequence typing (SLST) scheme. We evaluated the ability of the biofilm formation of C. acnes strains isolated from PJIs and 84 isolates recovered from healthy skin. Antibiotic susceptibility testing of planktonic and biofilm cells of PJI isolates and skin isolates was performed. Results: Most of the isolates from PJIs belonged to the SLST class H/phylotype IB (34.2%), followed by class D/phylotype IA1 (21.5%), class A/phylotype IA1 (18.9%), and class K/phylotype II (13.9%). All tested isolates were biofilm producers; no difference in biofilm formation was observed between the healthy skin group and the PJI group of strains. Planktonic and sessile cells of C. acnes remained highly susceptible to a broad spectrum of antibiotics, including beta-lactams, clindamycin, fluoroquinolones, linezolid, rifampin, and vancomycin. The minimal inhibitory concentrations (MICs) for planktonic and biofilm states coincided in most cases. However, the minimal biofilm eradication concentration (MBEC) was high for all antimicrobial drugs tested (>32 mg/L), except for rifampin (2 mg/L). Conclusions: C. acnes strains isolated from healthy skin were able to produce biofilm to the same extent as isolates recovered from PJIs. All C. acnes strains in planktonic and sessile states were susceptible to most antibiotics commonly used for PJI treatment, although rifampin was the only antimicrobial agent able to eradicate C. acnes embedded in biofilm.

Regulatory Networks Controlling Neurotoxin Synthesis in Clostridium botulinum and Clostridium tetani.

Clostridium botulinum and Clostridium tetani are Gram-positive, spore-forming, and anaerobic bacteria that produce the most potent neurotoxins, botulinum toxin (BoNT) and tetanus toxin (TeNT), responsible for flaccid and spastic paralysis, respectively. The main habitat of these toxigenic bacteria is the environment (soil, sediments, cadavers, decayed plants, intestinal content of healthy carrier animals). C. botulinum can grow and produce BoNT in food, leading to food-borne botulism, and in some circumstances, C. botulinum can colonize the intestinal tract and induce infant botulism or adult intestinal toxemia botulism. More rarely, C. botulinum colonizes wounds, whereas tetanus is always a result of wound contamination by C. tetani. The synthesis of neurotoxins is strictly regulated by complex regulatory networks. The highest levels of neurotoxins are produced at the end of the exponential growth and in the early stationary growth phase. Both microorganisms, except C. botulinum E, share an alternative sigma factor, BotR and TetR, respectively, the genes of which are located upstream of the neurotoxin genes. These factors are essential for neurotoxin gene expression. C. botulinum and C. tetani share also a two-component system (TCS) that negatively regulates neurotoxin synthesis, but each microorganism uses additional distinct sets of TCSs. Neurotoxin synthesis is interlocked with the general metabolism, and CodY, a master regulator of metabolism in Gram-positive bacteria, is involved in both clostridial species. The environmental and nutritional factors controlling neurotoxin synthesis are still poorly understood. The transition from amino acid to peptide metabolism seems to be an important factor. Moreover, a small non-coding RNA in C. tetani, and quorum-sensing systems in C. botulinum and possibly in C. tetani, also control toxin synthesis. However, both species use also distinct regulatory pathways; this reflects the adaptation of C. botulinum and C. tetani to different ecological niches.

Amplicon-Based Next-Generation Sequencing as a Diagnostic Tool for the Detection of Phylotypes of Cutibacterium acnes in Orthopedic Implant-Associated Infections.

The diagnosis of orthopedic implant-associated infections (OIAIs) caused by the slow-growing anaerobic bacterium Cutibacterium acnes is challenging. The mild clinical presentations of this low-virulent bacterium along with its ubiquitous presence on human skin and human-dominated environments often make it difficult to differentiate true infection from contamination. Previous studies have applied C. acnes phylotyping as a potential avenue to distinguish contamination from infection; several studies reported a prevalence of phylotypes IB [corresponding to type H in the single-locus sequence typing (SLST) scheme] and II (SLST type K) in OIAIs, while a few others found phylotype IA1 (more specifically SLST type A) to be abundant. However, phylotype determination has mainly been done in a culture-dependent manner on randomly selected C. acnes isolates. Here, we used a culture-independent amplicon-based next-generation sequencing (aNGS) approach to determine the presence and relative abundances of C. acnes phylotypes in clinical OIAI specimens. As amplicon, the SLST target was used, a genomic fragment that is present in all C. acnes strains known to date. The aNGS approach was applied to 30 sonication fluid (SF) samples obtained from implants removed during revision surgeries, including 17 C. acnes culture-positive and 13 culture-negative SF specimens. In 53% of the culture-positive samples, SLST types were identified: relative abundances were highest for K-type C. acnes, followed by H- and D-type C. acnes. Other types, including A- and C-type C. acnes that are more prevalent on human skin, had low relative abundances. The aNGS results were compared with, and confirmed by a culture-dependent approach, which included the isolation, whole genome sequencing (WGS) and phylotyping of 36 strains of C. acnes obtained from these SF samples. Besides serving as a powerful adjunct to identify C. acnes phylotypes, the aNGS approach could also distinguish mono- from heterotypic infections, i.e., infections caused by more than one phylotype of C. acnes: in eight out of nine culture-positive SF samples multiple C. acnes types were detected. We propose that the aNGS approach, along with the patient's clinical information, tissue and SF cultures and WGS, could help differentiate C. acnes contamination from true infection.

Solution Structure of the Cutibacterium acnes-Specific Protein RoxP and Insights Into Its Antioxidant Activity.

Cutibacterium acnes is a predominant bacterium on human skin and is generally regarded as commensal. Recently, the abundantly secreted protein produced by C. acnes, RoxP, was shown to alleviate radical-induced cell damage, presumably via antioxidant activity, which could potentially be harnessed to fortify skin barrier function. The aim of this study was to determine the structure of RoxP and elucidate the mechanisms behind its antioxidative effect. Here, we present the solution structure of RoxP revealing a compact immunoglobulin-like domain containing a long flexible loop which, in concert with the core domain, forms a positively charged groove that could function as a binding site for cofactors or substrates. Although RoxP shares structural features with cell-adhesion proteins, we show that it does not appear to be responsible for adhesion of C. acnes bacteria to human keratinocytes. We identify two tyrosine-containing stretches located in the flexible loop of RoxP, which appear to be responsible for the antioxidant activity of RoxP.

Diagnosis of orthopaedic-implant-associated infections caused by slow-growing Gram-positive anaerobic bacteria - a clinical perspective.

Slow-growing Gram-positive anaerobic bacteria (SGAB) such as Cutibacterium acnes are increasingly recognized as causative agents of implant-associated infections (IAIs) in orthopaedic surgeries. SGAB IAIs are difficult to diagnose because of their non-specific clinical and laboratory findings as well as the fastidious growth conditions required by these bacteria. A high degree of clinical suspicion and awareness of the various available diagnostic methods is therefore important. This review gives an overview of the current knowledge regarding SGAB IAI, providing details about clinical features and available diagnostic methodologies. In recent years, new methods for the diagnosis of IAI were developed, but there is limited knowledge about their usefulness in SGAB IAI. Further studies are required to determine the ideal diagnostic methodology to identify these infections so that they are not overlooked and mistakenly classified as aseptic failure.

Genomic Analysis of Cutibacterium acnes Strains Isolated from Prosthetic Joint Infections.

Cutibacterium acnes is a common cause of prosthetic joint infections (PJIs). The C. acnes population can be divided into six main phylotypes (IA1, IA2, IB, IC, II and III) that are associated with different clinical conditions and normal skin. A single-locus sequence typing (SLST) scheme can distinguish ten main SLST types: A-E (all IA1), F (IA2), G (IC), H (IB), K (II), L (III). We genome-sequenced and compared 16 strains of C. acnes isolated from healthy skin (n = 4) and PJIs (n = 12), including six PJI cases with a good outcome (four shoulder PJIs, one hip PJI, one knee PJI) and six with infection relapse (three shoulder PJIs, three hip PJIs). The sequenced strains belonged to four different phylotypes (IA1, IA2, IB and II) and seven different SLST types. All five type IB strains (all SLST type H1) were PJI isolates (three hip PJIs, two shoulder PJIs), and four of these caused infection relapse (three hip PJIs, one shoulder PJI). Isolates from PJI cases with a good outcome belonged to three different phylotypes (IA, IB, II). Interestingly, four strains (three strains from PJI cases with good outcome and one strain from healthy skin) contained a linear plasmid; these strains belonged to different SLST types (A1, C1, F4, H1) and were isolated in three different hospitals. This study suggests that type IB strains have the potential to cause infection relapse, in particular regarding hip PJIs. Moreover, our study revealed that strains belonging to the same SLST type can differ in their accessory genome in different geographic locations, indicative of microevolution.

Comparison of three amplicon sequencing approaches to determine staphylococcal populations on human skin.

BACKGROUND: Staphylococci are important members of the human skin microbiome. Many staphylococcal species and strains are commensals of the healthy skin microbiota, while few play essential roles in skin diseases such as atopic dermatitis. To study the involvement of staphylococci in health and disease, it is essential to determine staphylococcal populations in skin samples beyond the genus and species level. Culture-independent approaches such as amplicon next-generation sequencing (NGS) are time- and cost-effective options. However, their suitability depends on the power of resolution. RESULTS: Here we compare three amplicon NGS schemes that rely on different targets within the genes tuf and rpsK, designated tuf1, tuf2 and rpsK schemes. The schemes were tested on mock communities and on human skin samples. To obtain skin samples and build mock communities, skin swab samples of healthy volunteers were taken. In total, 254 staphylococcal strains were isolated and identified to the species level by MALDI-TOF mass spectrometry. A subset of ten strains belonging to different staphylococcal species were genome-sequenced. Two mock communities with nine and eighteen strains, respectively, as well as eight randomly selected skin samples were analysed with the three amplicon NGS methods. Our results imply that all three methods are suitable for species-level determination of staphylococcal populations. However, the novel tuf2-NGS scheme was superior in resolution power. It unambiguously allowed identification of Staphylococcus saccharolyticus and distinguish phylogenetically distinct clusters of Staphylococcus epidermidis. CONCLUSIONS: Powerful amplicon NGS approaches for the detection and relative quantification of staphylococci in human samples exist that can resolve populations to the species and, to some extent, to the subspecies level. Our study highlights strengths, weaknesses and pitfalls of three currently available amplicon NGS approaches to determine staphylococcal populations. Applied to the analysis of healthy and diseased skin, these approaches can be useful to attribute host-beneficial and -detrimental roles to skin-resident staphylococcal species and subspecies.

A Janus-Faced Bacterium: Host-Beneficial and -Detrimental Roles of Cutibacterium acnes.

The bacterial species Cutibacterium acnes (formerly known as Propionibacterium acnes) is tightly associated with humans. It is the dominant bacterium in sebaceous regions of the human skin, where it preferentially colonizes the pilosebaceous unit. Multiple strains of C. acnes that belong to phylogenetically distinct types can co-exist. In this review we summarize and discuss the current knowledge of C. acnes regarding bacterial properties and traits that allow host colonization and play major roles in host-bacterium interactions and also regarding the host responses that C. acnes can trigger. These responses can have beneficial or detrimental consequences for the host. In the first part of the review, we highlight and critically review disease associations of C. acnes, in particular acne vulgaris, implant-associated infections and native infections. Here, we also analyse the current evidence for a direct or indirect role of a C. acnes-related dysbiosis in disease development or progression, i.e., reduced C. acnes strain diversity and/or the predominance of a certain phylotype. In the second part of the review, we highlight historical and recent findings demonstrating beneficial aspects of colonization by C. acnes such as colonization resistance, immune system interactions, and oxidant protection, and discuss the molecular mechanisms behind these effects. This new insight led to efforts in skin microbiota manipulation, such as the use of C. acnes strains as probiotic options to treat skin disorders.

Prevalence of blaKPC-2, blaKPC-3 and blaKPC-30-Carrying Plasmids in Klebsiella pneumoniae Isolated in a Brazilian Hospital.

Klebsiella pneumoniae carbapenemase (KPC) actively hydrolyzes carbapenems, antibiotics often used a last-line treatment for multidrug-resistant bacteria. KPC clinical relevance resides in its widespread dissemination. In this work, we report the genomic context of KPC coding genes blaKPC-2, blaKPC-3 and blaKPC-30 in multidrug-resistant Klebsiellapneumoniae isolates from Brazil. Plasmids harboring blaKPC-3 and blaKPC-30 were identified. Fifteen additional carbapenem-resistant K. pneumoniae isolates were selected from the same tertiary hospital, collected over a period of 8 years. Their genomes were sequenced in order to evaluate the prevalence and dissemination of blaKPC-harboring plasmids. We found that blaKPC genes were mostly carried by one of two isoforms of transposon Tn4401 (Tn4401a or Tn4401b) that were predominantly located on plasmids highly similar to the previously described plasmid pKPC_FCF3SP (IncN). The identified pKPC_FCF3SP-like plasmids carried either blaKPC-2 or blaKPC-30. Two K. pneumoniae isolates harbored pKpQIL-like (IncFII) plasmids, only recently identified in Brazil; one of them harbored blaKPC-3 in a Tn4401a transposon. Underlining the risk of horizontal spread of KPC coding genes, this study reports the prevalence of blaKPC-2 and the recent spread of blaKPC-3, and blaKPC-30, in association with different isoforms of Tn4401, together with high synteny of plasmid backbones among isolates studied here and in comparison with previous reports.

Staphylococcus saccharolyticus Associated with Prosthetic Joint Infections: Clinical Features and Genomic Characteristics.

The anaerobic coagulase-negative staphylococcal species Staphylococcus saccharolyticus is a member of the normal skin microbiota. However, S. saccharolyticus is rarely found in clinical specimens and its pathogenic potential is unclear. The clinical data of prosthetic hip (n = 5) and shoulder (n = 2) joint implant-associated infections where S. saccharolyticus was detected in periprosthetic tissue specimens are described. The prosthetic hip joint infection cases presented as "aseptic" loosening and may represent chronic, insidious, low-grade prosthetic joint infections (PJIs), eventually resulting in loosening of prosthetic components. All cases were subjected to one-stage revision surgery and the long-term outcome was good. The shoulder joint infections had an acute onset. Polymicrobial growth, in all cases with Cutibacterium acnes, was found in 4/7 patients. All but one case were treated with long-term administration of beta-lactam antibiotics. Whole-genome sequencing (WGS) of the isolates was performed and potential virulence traits were identified. WGS could distinguish two phylogenetic clades (clades 1 and 2), which likely represent distinct subspecies of S. saccharolyticus. Little strain individuality was observed among strains from the same clade. Strains of clade 2 were exclusively associated with hip PJIs, whereas clade 1 strains originated from shoulder PJIs. It is possible that strains of the two clades colonize different skin habitats. In conclusion, S. saccharolyticus has the potential to cause PJIs that were previously regarded as aseptic loosening of prosthetic joint devices.

Complete Genome Sequences of Two Corynebacterium macginleyi Strains Isolated from Infectious Keratitis.

Corynebacterium macginleyi is a slow-growing, lipid-requiring bacterium that may cause ocular infections. Here, we report the complete genome sequences of two strains, T160811 and T180208, isolated from infectious keratitis. The two genomes consist of circular chromosomes of 2,431,961 bp and 2,481,998 bp, respectively, and contain high numbers of repetitive elements.