Chinese sacbrood virus (CSBV) is the major cause and lead to the collapse of Apis cerana colonies. VP1, the structural protein of CSBV, shows the highest variation in the amino acid sequences among proteins from different CSBV strains as well as exhibits excellent immunogenicity. However, its function with host protein still remains unclear. To clarify its function with host protein, we screened out host cellular proteins that interact with VP1 using the membrane protein yeast two-hybrid system. In addition, we verified interactions between heat shock protein 70 cognate 5 (Hsp70-c5) and VP1 using glutathione S-transferase (GST) pull-down and co-immunoprecipitation assays. VP1 and Hsp70-c5 were colocalized in the cytoplasm and nucleus. Using western blot and real-time polymerase chain reaction (PCR), Hsp70-c5 expression in CSBV-infected larvae was upregulated compared with that in healthy larvae. We observed that when we silenced Hsp70-c5, VP1 expression was significantly downregulated. These results demonstrate that Hsp70-c5 is involved in at least one stage(s) of the viral life cycle.

Despite being potentially useful extremophile resources, there have been few reports on acid-tolerant fungi and their bioactive metabolites. Acidophilic/aciduric fungi (n = 237) were isolated from Thai mangrove sediments in an acidic medium. Using fungal identification technology (including morphologic observation, chemical screening, and sequence comparisons) all the isolates were identified and 41 representative isolates were selected for analysis of the phylogenetic relationships (ITS rDNA, β-tubulin, calmodulin, and actin gene sequences). There were seven genera identified - Penicillium; Aspergillus; Talaromyces; Cladosporium; Allophoma; Alternaria; and Trichoderma - in four taxonomic orders of the phylum Ascomycota, and Penicillium, Aspergillus, and Talaromyces were the dominant genera. Acidity tolerance was evaluated and 95% of the isolates could grow under extremely acidic conditions (pH 2). Six strains were classed as acidophilic fungi that cannot survive under pH 7, all of which had an extraordinarily close genetic relationship and belonged to the genus Talaromyces. This is the first report on the acidophilic characteristics of this genus. The antimicrobial, anti-tumor, and antiviral activities of the fermentation extracts were evaluated. Nearly three-quarters of the extracts showed cytotoxic activity, while less than a quarter showed antimicrobial or anti-H1N1 activity. The typical aciduric fungus Penicillium oxalicum OUCMDZ-5207 showed similar growth but completely different chemical diversity at pH 3 and 7. The metabolites of OUCMDZ-5207 that were obtained only at pH 3 were identified as tetrahydroauroglaucin (1), flavoglaucin (2), and auroglaucin (3), among which auroglaucin showed strong selective inhibition of A549 cells with an IC50 value of 5.67 μM. These results suggest that acid stress can activate silent gene clusters to expand the diversity of secondary metabolites, and the bioprospecting of aciduric/acidophilic microorganism resources in Thai mangrove sediments may lead to the discovery of compounds with potential medicinal applications.

Non-tuberculous mycobacteria (NTM) are widespread in the environment and are a public health concern due to their resistance to antimicrobial agents. The colonization of surgical heater-cooler devices (HCDs) by the slow-growing NTM species Mycobacterium chimaera has recently been linked to multiple invasive infections in patients worldwide. The resistance of M. chimaera to antimicrobials may be aided by a protective biofilm matrix of extracellular polymeric substances (EPS). This study explored the hypothesis that M. chimaera can form biofilms on medically relevant materials. Several M. chimaera strains, including two HCD isolates, were used to inoculate a panel of medical device materials. M. chimaera colonization of the surfaces was monitored for 6 weeks. M. chimaera formed a robust biofilm at the air-liquid interface of borosilicate glass tubes, which increased in mass over time. M. chimaera was observed by 3D Laser Scanning Microscopy to have motility during colonization, and form biofilms on stainless steel, titanium, silicone and polystyrene surfaces during the first week of inoculation. Scanning electron microscopy (SEM) of M. chimaera biofilms after 4 weeks of inoculation showed that M. chimaera cells were enclosed entirely in extracellular material, while cryo-preserved SEM samples further revealed that an ultrastructural component of the EPS matrix was a tangled mesh of 3D fiber-like projections connecting cells. Considering that slow-growing M. chimaera typically has culture times on the order of weeks, the microscopically observed ability to rapidly colonize stainless steel and titanium surfaces in as little as 24 h after inoculation is uncharacteristic. The insights that this study provides into M. chimaera colonization and biofilm formation of medical device materials are a significant advance in our fundamental understanding of M. chimaera surface interactions and have important implications for research into novel antimicrobial materials, designs and other approaches to help reduce the risk of infection.

As one of the main pathogens of periodontitis, Porphyromonas gingivalis often forms mixed biofilms with other bacteria or fungi under the gingiva, such as Candida albicans. Heme is an important iron source for P. gingivalis and C. albicans that supports their growth in the host. From the perspective of heme competition, this study aims to clarify that the competition for heme enhances the pathogenic potential of P. gingivalis during the interaction between P. gingivalis and C. albicans. Porphyromonas gingivalis single-species biofilm and P. gingivalis-C. albicans dual-species biofilm were established in a low- and high-heme environment. The results showed that the vitality of P. gingivalis was increased in the dual-species biofilm under the condition of low heme, and the same trend was observed under a laser confocal microscope. Furthermore, the morphological changes in P. gingivalis were observed by electron microscope, and the resistance of P. gingivalis in dual-species biofilm was stronger against the killing effect of healthy human serum and antibiotics. The ability of P. gingivalis to agglutinate erythrocyte was also enhanced in dual-species biofilm. These changes disappeared when heme was sufficient, which confirmed that heme competition was the cause of thepathogenicy change in P. gingivalis. Gene level analysis showed that P. gingivalis was in a superior position in the competition relationship by increasing the expression of heme utilization-related genes, such as HmuY, HmuR, HusA, and Tlr. In addition, the expression of genes encoding gingipains (Kgp, RgpA/B) was also significantly increased. They not only participate in the process of utilizing heme, but also are important components of the virulence factors of P. gingivalis. In conclusion, our results indicated that the pathogenic potential of P. gingivalis was enhanced by C. albicans through heme competition, which ultimately promoted the occurrence and development of periodontitis and, therefore, C. albicans subgingival colonization should be considered as a factor in assessing the risk of periodontitis.

Phosphorus solubilizing bacteria (PSB) can promote the level of plant-absorbable phosphorus (P) in agro-ecosystems. However, little attention has been paid to PSB harboring abilities in utilizing multiple phosphorus sources and their potentials for heavy metal immobilization. In this study, we applied the strategy of stepwise acclimation by using Ca3(PO4)2, phytate, FePO4, and AlPO4 as sole P source. We gained 18 PSB possessing abilities of multiple P sources utilization, and these bacteria belonged to eight genera (Acinetobacter, Pseudomonas, Massilia, Bacillus, Arthrobacter, Stenotrophomonas, Ochrobactrum, and Cupriavidus), and clustered to two apparent parts: Gram-positive bacteria and Gram-negative bacteria. The isolate of Acinetobacter pittii gp-1 presented good performance for utilizing Ca3(PO4)2, FePO4, AlPO4, and phytate, with corresponding P solubilizing levels were 250.77, 46.10, 81.99, and 7.91 mg/L PO4 3--P, respectively. The PSB A. pittii gp-1 exhibited good performance for solubilizing tricalcium phosphate in soil incubation experiments, with the highest values of water soluble P and available P were 0.80 and 1.64 mg/L, respectively. Additionally, the addition of A. pittii gp-1 could promote the immobilization of lead (Pb), and the highest Pb immobilization efficiency reached 23%. Simultaneously, we found the increases in abundances of both alkaline phosphatase gene (phoD) and β-propeller phytase gene (bpp) in strain gp-1 added soils. Besides, we observed the expression up-regulation of both pyrroloquinoline quinone gene (pqq) and polyphosphate kinases gene (ppk), with the highest relative expression levels of 18.18 and 5.23, respectively. We also found the polyphosphate particles using granule staining. To our knowledge, our findings first suggest that the solubilizing of tricalcium phosphate by phosphorus solubilizing bacterium belonging to Acinetobacter is coupled with the synthesis of polyphosphate. Taken together, A. pittii gp-1 could be a good candidate in improving soil fertility and quality.

Lysine-porphyrin conjugate 4i has potent photosensitive antibacterial effect on clinical isolated bacterial strains such as Methicillin-resistant Staphylococcus aureus (MRSA), Escherichia coli, and Pseudomonas aeruginosa. The mechanism of photodynamic antibacterial chemotherapy of 4i (4i-PACT) in vitro and the treatment effect in vivo was investigated in this paper. Atomic force microscopy (AFM) revealed that 4i-PACT can effectively destroy membrane and wall of bacteria, resulting in leakage of its content. This was confirmed by dual fluorescent staining with acridine orange/ethidium bromide and measuring materials absorption at 260 nm. Agarose gel electrophoresis measurement showed that 4i-PACT can damage genomic DNA. Healing of wound in rat infected by mixed bacteria showed that the efficiency of 4i-PACT is dependent on the dose of light. These results showed that 4i-PACT has promising bactericidal effect both in vitro and in vivo.

The objective of this study was to evaluate the effects of wilting and Lactobacillus plantarum (LP) addition on the silage fermentation quality and microbial community of Moringa oleifera Lam. leaf silage. Unwilted (direct-cut) or wilted M. oleifera leaves were prepared either with or without LP (1.0 × 106 cfu/g) followed by either 60 or 120 days of ensiling, leading to eight treatment groups. The results showed that lactic acid was the dominant fermentation product, and no butyric acid was detected for any of the treatments. Higher acetic acid and propionic acid were detected during the fermentation of wilted silage compared to unwilted silage. Although NH3-N content increased after wilting, the content was far below 10% of the dry matter (DM). In addition, higher pH was observed after 120 days of ensiling compared to 60 days. Wilting also influenced the bacterial community structure. Lactobacillus was the most dominant genus in unwilted samples while Enterobacteriales, Weissella, and Pantoea were the most dominant genera in wilted samples. Furthermore, the relative abundance of undesirable microorganisms was far below that of lactic acid bacteria in all treatments. In summary, wilting had significant effects on fermentation quality, and it was shown that M. oleifera leaves can undergo quality ensiling directly without the addition of LP.

The report describes a simple, rapid and sensitive assay for visual and multiplex detection of Enterococcus faecalis and Staphylococcus aureus based on multiple loop-mediated isothermal amplification (mLAMP) and lateral flow biosensor (LFB). Detection and differentiation of the Ef0027 gene (E. faecalis-specific gene) and nuc gene (S. aureus-specific gene) were determined using fluorescein (FITC)-and digoxin-modified primers in the mLAMP process. In the presence of biotin- and FITC-/digoxin-modified primers, the mLAMP yielded numerous biotin- and FITC-/digoxin-attached duplex products, which were detected by LFB through biotin/streptavidin interaction (biotin on the duplex and streptavidin on the gold nanoparticle) and immunoreactions (FITC/digoxin on the duplex and anti-FITC/digoxin on the LFB test line). The accumulation of gold nanoparticles generated a characteristic red line, enabling visual and multiplex detection of target pathogens without instrumentation. The limit of detection (LoD), analytical specificity and feasibility of LAMP-LFB technique were successfully examined in pure culture and blood samples. The entire procedure, including specimen (blood samples) processing (30 min), isothermal reaction (40 min) and result reporting (within 2 min), could be completed within 75 min. Thus, this assay offers a simple, rapid, sensitive and specific test for multiplex detection of E. faecalis and S. aureus strains. Furthermore, the LAMP-LFB strategy is a universal technique, which can be extended to detect various target sequences by re-designing the specific LAMP primers.

Until now, there are few studies and reports on the use of endogenous promoters of obligate biotrophic fungi. The WY195 promoter in the genome of Oidium heveae, the rubber powdery mildew pathogen, was predicted using PromoterScan and its promoter function was verified by the transient expression of the β-glucuronidase (GUS) gene. WY195 drove high levels of GUS expression in dicotyledons and monocotyledons. qRT-PCR indicated that GUS expression regulated by the WY195 promoter was 17.54-fold greater than that obtained using the CaMV 35S promoter in dicotyledons (Nicotiana tabacum), and 5.09-fold greater than that obtained using the ACT1 promoter in monocotyledons (Oryza sativa). Furthermore, WY195-regulated GUS gene expression was induced under high-temperature and drought conditions. Soluble proteins extracted from WY195-hpaXm transgenic tobacco was bioactive. Defensive micro-HR induced by the transgene expression of hpaXm was observed on transgenic tobacco leaves. Disease resistance bioassays showed that WY195-hpaXm transgenic tobacco enhanced the resistance to tobacco mosaic virus (TMV). WY195 has great potential for development as a new tool for genetic engineering. Further in-depth studies will help to better understand the transcriptional regulation mechanisms and the pathogenic mechanisms of O. heveae.

Most isolated strains of Staphylococcus sciuri contain mecA1, the evolutionary origin of mecA, but are sensitive to β-lactams (OS-MRSS, oxacillin-susceptible mecA1-positive S. sciuri). In order to improve the efficacy of antibiotic treatment, it is important to clarify whether the resistance of OS-MRSS to β-lactams is an inducible phenotype. In this study, three OS-MRSS strains with oxacillin MIC = 1 μg/ml were isolated from 29 retail pork samples. The resistance of OS-MRSS to β-lactams (MIC > 256 μg/ml) was found to be induced by oxacillin, and the induced resistance was observed to remain stable within a certain period of time. Interestingly, the induced β-lactam resistance was not caused by mecA1, heterogeneous resistance, or any genetic mutation, but mainly due to increased wall teichoic acid (WTA) synthesis that thickened the cell wall. The induced strains also showed slower growth rate, as well as decreased adhesion ability and biofilm thickness. These phenotypes were found to be achieved through altered gene expression in associated pathways, such as the citrate cycle and pentose phosphate pathway. The results challenge the traditional antibiotic sensitivity test. In the presence of β-lactam antibiotics, OS-MRSS that was initially sensitive to β-lactams was observed to gradually develop β-lactam resistance in several days. This often-neglected phenomenon in antibiotic sensitivity tests requires further research attention.

Fungi are ubiquitous in nature; that is, they are present everywhere on the planet; understanding the active state and functional capacity of airborne microbes associated with health of human, animal, and plant is critical for biosafety management. Here, we firstly and directly proved that there were about 40% active fungi in the air via rRNA amplicon sequencing and imaging flow cytometry simultaneously. Amplicon sequencing analysis showed differences between structures of active and total fungal community; Ascomycota were dominant in the active community, while Basidiomycota have low transcriptional activity across all samples. Notably, plant pathogenic fungi were predominant in the air, and more than 50% were active, including not only several common plant pathogens but also biotrophic fungi (Erysiphe sp. and Microbotryum sp.) and host-specific pathogens, which were generally considered to be inactive after leaving the host. Putative plant pathogens of eight genera were found active across the sampling season, indicating their superior ability to obtain nutrients even in barren nutrient environments. Interestingly, we detected several potentially active unrecorded fungi in China (Diatrype prominens, Septofusidium herbarum, Pseudomicrostroma glucosiphilum, and Uromycladium tepperianum), which suggested that they spread over a long distance by air and may cause diseases under favorable conditions. Our results suggested that maintaining transmission in air is an essential feature of many fungi including plant pathogens regardless of being a biotrophic, hemibiotrophic, or necrotrophic group. Moreover, two potentially active human pathogens and one animal pathogen were captured, which indicated their potential risks. This study provided a new perspective for more comprehensive understanding of airborne fungi, including their multidimensional lifestyle, state, functioning, and potential pathogenic risk. It also laid the foundation for further prediction and management of airborne microbial communities, which would be of interest for public health and agriculture.

Virus infection has been consistently threatening public health. The cyclic GMP-AMP synthase (cGAS)-Stimulator of Interferon Genes (STING) pathway is a critical defender to sense various pathogens and trigger innate immunity of mammalian cells. cGAS recognizes the pathogenic DNA in the cytosol and then synthesizes 2'3'-cyclic GMP-AMP (2'3'cGAMP). As the second messenger, cGAMP activates STING and induces the following cascade to produce type I interferon (IFN-I) to protect against infections. However, viruses have evolved numerous strategies to hinder the cGAS-STING signal transduction, promoting their immune evasion. Here we outline the current status of the viral evasion mechanism underlying the regulation of the cGAS-STING pathway, focusing on how post-transcriptional modifications, viral proteins, and non-coding RNAs involve innate immunity during viral infection, attempting to inspire new targets discovery and uncover potential clinical antiviral treatments.

Listeria monocytogenes, an important food-borne pathogen, causes listeriosis and is widely distributed in many different environments. In a previous study, we developed a novel enrichment broth containing D-allose that allows better isolation of L. monocytogenes from samples. However, the mechanism of D-allose utilization by L. monocytogenes remains unclear. In the present study, we determined the metabolism of D-allose in L. monocytogenes and found that lineage II strains of L. monocytogenes can utilize D-allose as the sole carbon source for growth, but lineage I and III strains cannot. Transcriptome analysis and sequence alignment identified six genes (lmo0734 to 0739) possibly related to D-allose metabolism that are only present in the genomes of lineage II strains. Recombinant strain ICDC-LM188 containing these genes showed utilization of D-allose by growth assays and Biolog phenotype microarrays. Moreover, lmo0734 to 0736 were verified to be essential for D-allose metabolism, lmo0737 and 0738 affected the growth rate of L. monocytogenes in D-allose medium, while lmo0739 was dispensable in the metabolism of D-allose in L. monocytogenes. This is the first study to identify the genes related to D-allose metabolism in L. monocytogenes, and their distribution in lineage II strains. Our study preliminarily determined the effects of these genes on the growth of L. monocytogenes, which will benefit the isolation and epidemiological research of L. monocytogenes.

Staphylococcus aureus (S. aureus), including methicillin-resistant S. aureus (MRSA), is one of the most important human pathogens, which is responsible for bacteremia, soft-tissue infections, and food poisoning. Hence, multiple cross displacement amplification (MCDA) is employed to detect all S. aureus strains, and differentiates MRSA from methicillin-sensitive S. aureus. Multiplex MCDA (m-MCDA), which targets the nuc gene (S. aureus-specific gene) and mecA gene (encoding penicillin-binding protein-2'), could detect S. aureus strains and identify MRSA within 85 min. Detection of the m-MCDA products is achieved using disposable lateral flow biosensors. A total of 58 strains, including various species of Gram-positive and Gram-negative strains, are used for evaluating and optimizing m-MCDA assays. The optimal amplification condition is found to be 63°C for 40 min, with detection limits at 100 fg DNA/reaction for nuc and mecA genes in the pure cultures, and 10 CFU/tube for nuc and mecA genes in the blood samples. The analytical specificity of m-MCDA assay is of 100%, and no cross-reactions to non-S. aureus strains are produced according to the specificity testing. Particularly, two additional components, including AUDG enzyme and dUTP, are added into the m-MCDA amplification mixtures, which are used for eliminating the unwanted results arising from carryover contamination. Thus, the m-MCDA technique appears to be a simple, rapid, sensitive, and reliable assay to detect all S. aureus strains, and identify MRSA infection for appropriate antibiotic therapy.

Our previous study found that LeDnaJ07 RNAi decreased Lentinula edodes resistance to heat stress and Trichoderma atroviride infection. In this study, the structure and function of the LeDnaJ07 gene was analyzed by gene cloning and overexpression in L. edodes stress-sensitive strain YS55 via the Agrobacterium-mediated transformation method. Transformants were confirmed by qRT-PCR, fluorescence observation and Southern blotting. Overexpression of LeDnaJ07 in YS55 not only enhanced L. edodes mycelial resistance to heat stress but also facilitated mycelial growth. In the presence of heat stress, the intracellular IAA content showed a significant increase in the two LeDnaJ07 overexpression strains but only a slight change in the YS55 wild type strain. Moreover, the interaction between LeDnaJ07 and LetrpE was demonstrated via Y2H and BiFC assays. These results suggested that LeDnaJ07 may be involved in regulating IAA biosynthesis and the resistance of L. edodes to heat stresses via interacting with LetrpE.

Microbial-based therapies are one of the hotspots in the field of ulcerative colitis research. The lactic acid bacteria and their postbiotics occupy a key position in microbial therapies, however, the mechanism by which they alleviate ulcerative colitis in mice is unknown. We investigated the effects of Lacticaseibacillus rhamnosus 2016SWU.05.0601 (Lr-0601) and its postbiotics on male Kunming mice with dextran sulfate sodium salt (DSS)-induced ulcerative colitis (UC). The results showed that Lr-0601 significantly alleviated the deterioration of UC and restored the expression of intestinal mechanical barrier proteins. In addition, Lr-0601 significantly reduced the expression of inflammatory cytokines in the body and regulated the expression of key regulatory genes of the NF-κB-iNOS/COX-2 signaling pathway in colon tissues to a large extent. Our results suggest that supplementation with Lr-0601 and its postbiotics can effectively prevent DSS-induced UC and have a beneficial effect on intestinal health, which also provides new insights and research bases for the prevention as well as the treatment of ulcerative colitis and other diseases related to intestinal barrier dysfunction and other diseases.

The arginine catabolic mobile element (ACME) facilitates colonization of staphylococci on skin and mucous membranes by improving their tolerances to polyamines and acidic conditions. ACME is inserted in tandem with the SCCmec element and Staphylococcus epidermidis has been proposed to be a reservoir of ACME for other staphylococci. In this study, we investigated the existence of ACME in 146 staphylococcal isolates from mastitic milk and found 21 of them carried ACME. Almost half of the investigated S. epidermidis isolates contained the element. The whole genome of a S. epidermidis strain Y24 with ACME was further sequenced and the ACME-SCCmec composite island was assembled. This composite island is 81.3 kb long and consisted of 77 ORFs including a methicillin resistance gene mecA, a type II' ACME gene cluster, a virulence gene pls and eight heavy metal tolerance genes. Wide existence of ACME in livestock-associated staphylococci from this study and a potential risk of spreading ACME among different staphylococcal species warrant close monitoring and further studies.

Septoria nodorum blotch is a major disease of wheat caused by the fungus Parastagonospora nodorum. Recent studies have demonstrated that secondary metabolites, including polyketides and non-ribosomal peptides, produced by the pathogen play important roles in disease and development. However, there is currently no knowledge on the composition or biological activity of the volatile organic compounds (VOCs) secreted by P. nodorum. To address this, we undertook a series of growth and phytotoxicity assays and demonstrated that P. nodorum VOCs inhibited bacterial growth, were phytotoxic and suppressed self-growth. Mass spectrometry analysis revealed that 3-methyl-1-butanol, 2-methyl-1-butanol, 2-methyl-1-propanol, and 2-phenylethanol were dominant in the VOC mixture and phenotypic assays using these short chain alcohols confirmed that they were phytotoxic. Further analysis of the VOCs also identified the presence of multiple sesquiterpenes of which four were identified via mass spectrometry and nuclear magnetic resonance as β-elemene, α-cyperone, eudesma-4,11-diene and acora-4,9-diene. Subsequent reverse genetics studies were able to link these molecules to corresponding sesquiterpene synthases in the P. nodorum genome. However, despite extensive testing, these molecules were not involved in either of the growth inhibition or phytotoxicity phenotypes previously observed. Plant assays using mutants of the pathogen lacking the synthetic genes revealed that the identified sesquiterpenes were not required for disease formation on wheat leaves. Collectively, these data have significantly extended our knowledge of the VOCs in fungi and provided the basis for further dissecting the roles of sesquiterpenes in plant disease.

Shigella spp., the etiological agent of shigellosis or "bacillary dysentery," are responsible for considerable morbidity and mortality in excess of a million deaths globally per year. Although PCR-based techniques (such as PCR-based dipstick biosensors) have been used for the molecular diagnosis of infectious disease, these assays were restricted due to the need for a sophisticated thermal cycling apparatus to denature target templates. To facilitate simple and rapid detection of target pathogens, we successfully devised an inexpensive, reliable and nearly instrument-free molecular technique, which incorporates multiple cross displacement amplification (MCDA) combined with a newly designed lateral flow biosensor (LFB) for visual, sensitive and specific detection of Shigella. The MCDA-LFB assay was conducted at 65°C for only 20 min during the amplification stage, and then products were directly analyzed on the biosensor, alleviating the use of special reagents, electrophoresis equipment and amplicon detection instruments. The entire process, including specimen processing (35 min), amplification (20) and detection (2-5 min), can be finished within 1 h. The MCDA-LFB assay demonstrated high specificity for Shigella detection. The analytical sensitivity of the assay was 10 fg of genomic templates per reaction in pure culture and 5.86 CFU per tube in human fecal samples, which was consistent with MCDA by colorimetric indicator, gel electrophoresis, real time turbidity and fluorescence detection. Hence, the simplicity, rapidity and nearly instrument-free platform of the MCDA-LFB assay make it practical for 'on-site' diagnosis, point-of-care testing and more. Moreover, the proof-of-concept approach can be reconfigured to detect a wide variety of target sequences by re-designing the specific MCDA primers.

Urinary tract infections (UTIs) are one of the most common bacterial infections in humans, causing cystitis, pyelonephritis, and renal failure. Uropathogenic Escherichia coli (UPEC) is the leading cause of UTIs. Accurate and rapid discrimination of UPEC lineages is useful for epidemiological surveillance. Fimbriae are necessary for the adherence of UPEC strains to host uroepithelia, and seem to be abundant and diverse in UPEC strains. By analyzing all the possible fimbrial operons in UPEC strains, we found that closely related strains had similar types of chaperone-usher fimbriae, and the diversity of fimbrial genes was higher than that of multilocus sequence typing (MLST) genes. A typing assay based on the polymorphism of four gene sequences (three fimbrial genes and one housekeeping gene) and the diversity of fimbriae present was developed. By comparison with the MLST, whole-genome sequence (WGS) and fumC/fimH typing methods, this was shown to be accurate and have high resolution, and it was also relatively inexpensive and easy to perform. The assay can supply more discriminatory information for UPEC lineages, and have the potential to be applied in epidemiological surveillance of UPEC isolates.