Isolation and characterization of bacteria associated with silkworm gut under antibiotic-treated larval feeding / Isolamento e caracterização de bactérias associadas ao trato intestinal do bicho-da-seda sob alimentação larval tratada com antibióticos

Abstract The impact of antibiotics on growth, cocoon production was assessed in addition to isolation and characterization of bacteria associated with silkworm gut of infected larvae. Larval rearing was maintained at recommended conditions of temperature and humidity. Silkworm larvae showing abnormal symptoms were collected from the control group and dissected for gut collection. Bacteria were isolated from the gut content by spreading on agar plates and incubated at 37 °C for 48 hrs. Bacterial identification and phylogenetic analysis were carried out by 16S rRNA gene sequencing. The isolated bacteria were subjected to antimicrobial susceptibility test (disc diffusion methods) by using Penicillin (10 µg/mL), Tetracycline (30 µg/mL), Amoxicillin (25 µg/mL), Ampicillin (10 µg/mL), and Erythromycin (15 µg/mL). All isolated strains showed positive results for the catalase test. We isolated and identified bacterial strains (n = 06) from the gut of healthy and diseased silkworm larvae. Based on the 16S rRNA gene sequence, isolated bacteria showed close relation with Serratia, Bacillus, and Pseudomonas spp. Notably, 83.3% of strains were resistant to Penicillin, Tetracycline, Amoxicillin, Ampicillin, and Erythromycin but 16.6% showed antibiotic susceptibility to the above-mentioned commonly used antibiotics. Silkworm larvae fed on penicillin-treated leaves showed significant improvement in larval weight, larval length, and cocoon production. Significantly higher larval weight (6.88g), larval length (5.84cm), and cocoon weight (1.33g) were recorded for larvae fed on leaves treated with penicillin as compared to other antibiotics. Isolated bacterial strains showed close relation with Serratia spp., Bacillus spp. and Pseudomonas spp.

Resumo O impacto dos antibióticos no crescimento e na produção do casulo foi avaliado, além do isolamento e caracterização das bactérias associadas ao intestino de larvas infectadas do bicho-da-seda. A criação das larvas foi mantida nas condições recomendadas de temperatura e umidade. As larvas do bicho-da-seda com sintomas anormais foram coletadas do grupo controle e dissecadas para coleta do intestino. As bactérias foram isoladas do conteúdo intestinal por espalhamento em placas de ágar e incubadas a 37° C durante 48 horas. A identificação bacteriana e a análise filogenética foram realizadas pelo sequenciamento do gene 16S rRNA. As bactérias isoladas foram submetidas a teste de sensibilidade antimicrobiana (métodos de difusão em disco) com penicilina (10 µg / mL), tetraciclina (30 µg / mL), amoxicilina (25 µg / mL), ampicilina (10 µg / mL) e eritromicina (15 µg / mL). Todas as cepas isoladas apresentaram resultados positivos para o teste da catalase. Isolamos e identificamos cepas bacterianas (n = 06) do intestino de larvas de bicho-da-seda saudáveis e doentes. Com base na sequência do gene 16S rRNA, as bactérias isoladas mostraram estreita relação com Serratia, Bacillus e Pseudomonas spp. Notavelmente, 83,3% das cepas eram resistentes a penicilina, tetraciclina, amoxicilina, ampicilina e eritromicina, mas 16,6% mostraram suscetibilidade aos antibióticos comumente usados mencionados acima. As larvas do bicho-da-seda alimentadas com folhas tratadas com penicilina apresentaram melhora significativa no peso larval, comprimento larval e produção de casulo. Peso larval significativamente maior (6,88g), comprimento larval (5,84cm) e peso do casulo (1,33g) foram registrados para larvas alimentadas com folhas tratadas com penicilina, em comparação com outros antibióticos. Cepas bacterianas isoladas mostraram estreita relação com Serratia spp., Bacillus spp. e Pseudomonas spp.

Percentage Incidences of Bacteria in Mahseer (Tor putitora), Silver carp (Hypophthalmichthys molitrix) Fish Collected from Hatcheries and Local Markets of District Malakand and Peshawar of Khyber Pakhtunkhwa, Pakistan / Incidências percentuais de bactérias em Mahseer (Tor putitora), carpa-prateada (Hypophthalmichthys molitrix), peixes coletados em incubatórios e mercados locais do distrito de Malakand e Peshawar de Khyber Pakhtunkhwa, Paquistão

Abstract Fish is the main source of animal protein for human diet. The aim of this study was to find out prevalence of pathogenic bacteria of two selected economically important fish of Pakistan namely Mahseer (Tor putitora) and Silver carp (Hypophthalmichthys molitrix). Live fish samples from hatcheries and dead fish samples from different markets of study area were randomly collected. The fish samples were analyzed for isolation, identification and prevalence of bacteria. The isolated bacteria from study fish were identified through biochemical test and about 10 species of pathogenic bacteria were identified including the pathogenic bacteria to human and fish namely, Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus iniae, Serratia spp. Citrobacter spp. Stenotrophomonas spp. Bacillus spp. and Salmonella spp. The bacterial percentage frequency of occurrence in Silver carp and Mahseer fish showed Pseudomonas aeruginosa 21.42%, Staphylococcus epidermidis 17.85%, Escherichia coli 11.90%, Staphylococcus aureus 9.52%, Citrobacter spp. 9.52%, Serratia spp. 8.33%, Streptococcus iniae 7.14%, Stenotrophomonas spp. 5.95%, Bacillus spp. 4.76% and Salmonella spp. 3.57%. The study revealed that Fish samples of Mahseer and Silver carp that were collected from markets have found more isolates (10 bacterial species) than did the fresh fish pond samples (03 bacterial species) of hatcheries. The occurrence of pathogenic bacteria in study fish showed risk factor for public health consumers.

Resumo O peixe é a principal fonte de proteína animal para a alimentação humana. O objetivo deste estudo foi descobrir a prevalência de bactérias patogênicas de dois peixes economicamente importantes selecionados do Paquistão, nomeadamente Mahseer (Tor putitora) e carpa prateada (Hypophthalmichthys molitrix). Amostras de peixes vivos de incubatórios e amostras de peixes mortos de diferentes mercados da área de estudo foram coletadas aleatoriamente. As amostras de peixes foram analisadas quanto ao isolamento, identificação e prevalência de bactérias. As bactérias isoladas dos peixes do estudo foram identificadas através de testes bioquímicos e cerca de 10 espécies de bactérias patogênicas foram identificadas incluindo as bactérias patogênicas para humanos e peixes, nomeadamente, Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus iniae, Serratia spp. Citrobacter spp. Stenotrophomonas spp. Bacillus spp. e Salmonella spp. A porcentagem de freqüência de ocorrência bacteriana em carpa prateada e peixes Mahseer mostrou Pseudomonas aeruginosa 21,42%, Staphylococcus epidermidis 17,85%, Escherichia coli 11,90%, Staphylococcus aureus 9,52%, Citrobacter spp. 9,52%, Serratia spp. 8,33%, Streptococcus iniae 7,14%, Stenotrophomonas spp. 5,95%, Bacillus spp. 4,76% e Salmonella spp. 3,57%. O estudo revelou que as amostras de peixes de Mahseer e carpa prateada coletadas nos mercados encontraram mais isolados (10 espécies bacterianas) do que as amostras de peixes frescos (03 espécies bacterianas) de incubatórios. A ocorrência de bactérias patogênicas nos peixes do estudo apresentou fator de risco para consumidores de saúde pública.

Comparative in vitro analysis of the antifungal activity of different calcium silicate-based endodontic sealers

Aim: This study aimed to perform an in vitro comparative analysis of the antifungal activity of different calcium silicate-based endodontic sealers against three fungal species. Methods: The antifungal properties of three calcium silicate-based sealers were tested: Bio-C Sealer, Cambiar a Sealer Plus BC, and MTA-Fillapex. Two commonly used sealers were used as controls: AH Plus and Endomethasone. An agar diffusion test was performed to analyze the antifungal activity of the sealers against Candida albicans, Candida glabrata, Candida tropicalis, and a mixed microbial culture medium. The results were analyzed using ANOVA (p <0.05). Results: Endomethasone exhibited the highest inhibition against all strains examined, maintaining a consistent level of inhibition throughout 7 days. MTA-Fillapex demonstrated the best performance among the calcium silicate-based sealers for the three fungal species (p < 0.05), maintaining stable values over the 7 days, surpassing that of Endomethasone. Nevertheless, MTA-Fillapex only exhibited antimicrobial effect against the mixed culture for the first 24 hours, and no antimicrobial activity was observed at 48 hours, being surpassed by all tested sealers (p < 0.05). Conclusion: Of all silicate-based sealers tested, only MTA-Fillapex exhibited promising antifungal activity. Nevertheless, care must be taken when extrapolating these results, as MTA-Fillapex exhibited poor antimicrobial activity when tested in mixed microbial cultures

Mobile phones of anesthesiologists as reservoirs of nosocomial bacteria in a quaternary teaching hospital: an observational study

Abstract Background: Mobile phones in hospital settings have been identified as an important source of cross-contamination because of the low frequency with which mobile phones are cleaned by health workers and cyclical contamination of the hands and face. The aim of this study was to investigate whether the mobile phones of the anesthesia team at a teaching hospital are potential reservoirs of nosocomial bacteria. In addition, differences in device sanitization and hand hygiene habits between attending and resident anesthesiologists were correlated with mobile phone colonization. Methods: A prevalence study was conducted over a 6-month period from 2017 to 2018 that involved the collection of samples from the mobile phones of the anesthesiology team and culturing for surveillance. A questionnaire was administered to assess the mobile phone sanitization and hand washing routines of the anesthesia team in specific situations. Results: Bacterial contamination was detected for 86 of the 128 mobile phones examined (67.2%). A greater presence of Micrococcus spp. on devices was correlated with a higher frequency of mobile phone use (p = 0.003) and a lower frequency of sanitization (p = 0.003). The presence of bacteria was increased on the mobile phones of professionals who did not perform handwashing after tracheal intubation (p = 0.003). Conclusion: Hand hygiene and device sanitization habits were more important than the use behavior, as a higher presence of bacteria correlated with poorer hygiene habits. Furthermore, handwashing is the best approach to prevent serious colonization of mobile devices and the possible transmission of pathogens to patients under the care of anesthesiologists.

Helicobacter pylori: Microorganismo patógeno o mutualista en poblaciones colombianas / Helicobacter pylori: Pathogenic or mutualistic microorganism in Colombian populations / Helicobacter pylori: microrganismo patógeno ou mutualista em populações Colombianas

Introducción: El riesgo de desarrollar cáncer gástrico varía entre continentes, países y regiones. A pesar de que existe una alta prevalencia de Helicobacter pylori su rol como patógeno o mutualista define el riesgo de cáncer gástrico en las regiones de Colombia. Objetivo: Discutir el rol de Helicobacter pylori en el riesgo de cáncer gástrico en Colombia. Materiales y métodos: Revisión de literatura mediante la búsqueda, en las bases de datos LILACS, SciELO, PubMed. Resultados: La coevolución del humano y de Helicobacter pylori; la virulencia de genes cagA, vacA; el tipo de respuesta inmune inflamatoria a Helicobacter pylori (Th1) o antinflamatoria (Th2) y la susceptibilidad humana a cáncer gástrico (IL1β, IL10), junto a la dieta y factores ambientales explican el papel de Helicobacter pylori como patógeno o mutualista asociado al riesgo de cáncer gástrico en Colombia. Conclusiones: Helicobacter pylori tiene un rol mutualista principalmente en poblaciones de bajo riesgo de cáncer gástrico (costas), no obstante, en poblaciones con alto riesgo de cáncer gástrico (andes), su papel como patógeno amerita la erradicación; única estrategia para mitigar la alta incidencia de este cáncer en Colombia.

Introduction: The risk to develop gastric cancer varies between continents, countries and regions. Although there is a high prevalence of Helicobater pylori, its role as either pathogen or mutualistic bacteria defines the risk of gastric cancer in Colombian regions. Objective: To discuss the role of Helicobacter pylori in the risk of gastric cancer in Colombia. Materials and methods: A literature review based on searching LILACS, SciELO, and PubMed databases. Results: Helicobacter pylori role as either a pathogen or mutualistic microorganism associated with gastric cancer risk in Colombia can be explained by analyzing elements such as: human and Helicobacter pylori coevolution; cagA and vacA gene virulence; inflammatory (Th1) or anti-inflammatory (Th2) responses induced by Helicobacter pylori; human susceptibility to gastric cancer (IL1β, IL10); diet; and environmental factors. Conclusions: Even though Helicobacter pylori has a mutualistic role in populations at low gastric cancer risk (coastal regions), its role as a pathogen in populations at higher risk (Andean regions) justifies its eradication as a key strategy to mitigate the incidence of this cancer in Colombia.

Introdução: O risco de desenvolver câncer gástrico varia entre continentes, países e regiões. Embora haja uma alta prevalência de Helicobacter pylori, seu papel como patógeno ou mutualista define o risco de câncer gástrico nas regiões da Colômbia. Objetivo: Discutir o papel do Helicobacter pylori no risco de câncer gástrico na Colômbia. Materiais e métodos: Revisão da literatura por meio da busca, nas bases de dados LILACS, SciELO e PubMed. Resultados: A coevolução de humanos e Helicobacter pylori; a virulência dos genes cagA, vacA; o tipo de resposta imune inflamatória ao Helicobacter pylori (Th1) ou anti-inflamatório (Th2) e a suscetibilidade humana ao câncer gástrico (IL1β, IL10), juntamente com a dieta e fatores ambientais explicam o papel do Helicobacter pylori como patógeno ou mutualista associado ao risco de câncer gástrico na Colômbia. Conclusões: Helicobacter pylori tem um papel mutualista principalmente em populações de baixo risco de câncer gástrico (litoral), porém, em populações com alto risco de câncer gástrico (andes), seu papel como patógeno justifica a erradicação; única estratégia para mitigar a alta incidência deste câncer na Colômbia.

Cultivo versus metagenómica para la identificación bacteriana en pacientes con osteomielitis de pie diabético: una revisión sistemática / Culture and metagenomics as bacterial identification methods in patients with diabetic foot ulcers: a systematic review

BACKGROUND: Diabetic foot osteomyelitis (DFO) is a serious complication of infected ulcers in a diabetic patient. The identification of the infecting microorganisms is generally by culture, which causes a bias. Recently, metagenomics has been used for microbial identification. AIM: To systematically review the scientific literature related to DFO in the last 10 years to evaluate if culture and metagenomics are complementary. MATERIAL AND METHODS: To carry out the systematic review, PRISMA and Rayyan were used for the selection of studies, using three databases, using the keywords diabetes, osteomyelitis, culture and microbiome. Articles in English or Spanish were included, containing information related to bacterial identification in DFO. Characteristics of the technique, patients and frequency of bacterial appearance were collected. RESULTS: Twenty six articles were included, 19 used culture and 7 metagenomics. The patients were predominantly men (68%), with an average age of 61 years, 83% had type 2 diabetes and comorbidities, mainly vascular and neuropathy. The Families with the highest frequency of appearance using the culture technique were Enterobacteriaceae (29.3%) and Staphylococcaceae(28.3%) and with metagenomics Peptoniphilaceae (22.1%) and Staphylococcaceae (9.4%). Peptoniphilaceae were not identified in culture, although they were frequently identified by metagenomics. Methicillin- resistant Staphylococcus aureus, regularly identified by culture, was not identified using metagenomics. CONCLUSIONS: Comparing results, there is a certain complementarity between microbiological culture and sequencing to identify bacteria present in DFO.

Actividad antimicrobiana de extractos de plantas frente a Staphylococcus aislados de pacientes con conjuntivitis bacteriana / Antimicrobial Activity of Plant Extracts against Isolated Staphylococcus Infection in Patients with Bacterial Conjunctivitis / Atividade antimicrobiana de extratos vegetais contra Staphylococcus isolados de pacientes com conjuntivite bacteriana

Staphylococcus aureus y Staphylococcus epidermidis son los principales agentes etiológicos de las conjuntivitis bacterianas, que al tratarse con antibióticos de manera empírica, incrementan la resistencia antimicrobiana después de exposiciones repetidas. Se están buscando alternativas naturales para el tratamiento de infecciones bacterianas autolimitadas de la conjuntiva. Objetivo: determinar la actividad antimicrobiana de ocho extractos de las plantas frente a bacterias aisladas de pacientes con conjuntivitis bacterianas. Materiales y métodos: se tomaron muestras de 15 pacientes con conjuntivitis bacterianas. Se cultivaron en agar sangre y chocolate durante 24 h a 37 °C y se identificaron mediante el sistema automatizado vitek y pruebas de susceptibilidad antimicrobiana por el método de Kirby-Bauer. A cada aislamiento identificado con el género Staphylococcus se le evaluó su susceptibilidad frente a siete extractos: Ocimum basilicum, Sambucus nigra L., Delphinium elatum, Calendula officinalis, Bixa ore-llana (parte aérea y fruto independiente), Clinopodium brownei y Laurus nobilis, con un uso tradicional reportado para el tratamiento de infecciones oculares. Resultados: las bacterias aisladas con más frecuencia fueron S. epidermidis, S. hominis y S. aureus, las cuales presentaron resistencia antimicrobiana a oxacilina, tetraciclinas y eritromicina. Todos los aislamientos fueron inhibidos por los extractos de O. basilicum (cmi: >0.9 mg/mL) y L. nobilis (cmi: hasta 15 mg/mL). Conclusión: los extractos de C. officinalis y D. elatum tuvieron actividad antimicrobiana solo frente a los aislados con mayor sensibilidad antimi-crobiana. Los extractos etanólicos de O. basilicum y L. nobilis pueden ser una alternativa de tratamiento de las infecciones de la conjuntiva.

Staphylococcus aureus and Staphylococcus epidermidis are the primary etiological agents of bacterial conjunctivitis which are empirically treated with antibiotics. This results in an increase in antimicrobial resistance due to repeated exposure. Currently, natural treatment alternatives are being sought for self-limited bacterial infections of the conjunctiva. Objective: To determine the antimicrobial activity of eight extracts from Colombian plants against bacteria isolated from patients with bacterial conjunctivitis. Materials and methods: Samples were taken from 15 patients with bacterial conjunctivitis which were grown on blood and chocolate agar for 24 h at 37 °C. These samples were identified by the vitek automated system and antimicrobial susceptibility tests by the Kirby Bauer method. Each isolate identified with the genus Staphylococcus was evaluated for susceptibility to the following eight plant extracts of seven plant: Ocimum basilicum (basil), Sambucus nigra L. (elderberry), Delphinium elatum(belladonna), Calendula officinalis (marigold), Bixa orellana (annatto) (aerial part and independent fruit), Clinopodium brownei (pennyroyal), and Laurus nobilis (laurel), with traditional use previously reported for treating eye infections. Results: The most frequently isolated bacteria were S. epidermidis, S. hominis, and S. aureus, which exhibited antimicrobial resistance mainly to oxacillin, tetracyclines, and erythromycin. All isolates were inhibited by O. basilicum extracts (mic > 0.9 mg/mL) and L. nobilis (mic < 15 mg/mL). Conclusion: The extracts of C. officinalis y D. elatum showed antimicrobial activity only against isolates with higher antimicrobial sensitivity. Ethanolic extracts of O. basilicum y L. nobilis can be used as an alternative treatment for infections of the anterior segment of the eye.

Staphylococcus aureus e Staphylococcus epidermidis são os principais agentes etiológicos da conjuntivite bacteriana, estes são tratados empiricamente com antibióticos, causando aumento da resistência antimicrobiana após repetidas exposições aos mesmos. Atualmente, estão sendo estudadas alternativas naturais para o tratamento de infecções bacterianas autolimitadas da conjuntiva. Objetivo: determinar a atividade antimicrobiana de oito extratos de sete vegetais contra bactérias isoladas de pacientes com conjuntivite bacteriana. Materiais e métodos: foram retiradas amostras de 15 pacientes com conjuntivite bacteriana. As amostras foram cultivadas em ágar sangue e ágar chocolate por 24 horas a 37°C e os isolados foram identificados pelo sistema automatizado vitek, além de testes de susce-tibilidade antimicrobiana pelo método Kirby Bauer. Cada isolado identificado como sendo pertencente ao gênero Staphylococcus foi avaliado quanto à suscetibilidade a oito extratos vegetais: Ocimum basili-cum (manjericão), Sambucus nigra L. (sabugueiro), Delphinium elatum (belladona), Calendula officinalis(calêndula), Bixa orellana (urucum; parte aérea e fruto independente), Clinopodium brownei (poejo) e Laurus nobilis (louro), anteriormente relatados como uso tradicional para o tratamento de infecções ocu-lares. Resultados: as bactérias mais frequentemente isoladas foram S. epidermidis, S. hominis e S. aureus, que apresentaram resistência antimicrobiana principalmente à oxacilina, tetraciclinas e eritromicina. Todos os isolados foram inibidos por extratos de O. basilicum (cim: >0,9 mg/mL) e L. nobilis (cim: até 15 mg/mL). Conclusão: os extratos de C. officinalis e D. elatum apresentaram atividade antimicrobiana apenas contra os isolados com maior sensibilidade antimicrobiana. Os extratos etanólicos de O. basilicum e L. nobilis podem ser uma alternativa de tratamento para infecções conjuntivais.

Structural characteristics of lower respiratory tract microflora in patients with pneumoconiosis / 中华劳动卫生职业病杂志

Objective: To explore the composition of bacteria in lower respiratory tract of patients with pneumoconiosis and dust exposure, and to compare and analyze the difference and correlation between them. Methods: From May 2020 to January 2021, a prospective multicenter cross-sectional study was conducted to select patients with pneumoconiosis who underwent bronchoalveolar lavage treatment at the Respiratory and Critical Care Medical Department of the 920th Hospital of the Joint Support Force and the Respiratory Department of Tongren Hospital in Kunming, as well as the population of dust recipients. A total of 24 patients with pneumoconiosis (pneumoconiosis group) were included, and 16 dust exposed individuals (dust exposed group) were used as controls. Two groups of patients' alveolar lavage fluid were collected. The 16SrRNA gene V3-V4 sequencing technology and bioinformatics analysis platform were used to measure and analyze the differences in microbial structure composition and associations between bacterial communities. Results: Compared with the dust exposed group, the top 5 bacterial phyla in the alveolar lavage fluid level of patients with pneumoconiosis were the same, followed by Proteobacteria, Firmicutes, Bacteroidetes, Fusobacteria, and Actinobacteria. Compared with the dust exposure group, the pneumoconiosis group patients belong to the top 5 genera of horizontal flora abundance, which are different. The dust exposure group is respectively: Pseudomonas, Proctor, Streptococcus, Achromobacter, and Neisseria. The pneumoconiosis group is respectively: Pseudomonas, Achromobacter, Streptococcus, Ralstonia, and Proctor. The Alpha diversity analysis results showed that compared with the dust exposed group, the level of bacterial diversity in the pneumoconiosis group was difference (P<0.05), and there was no statistically significant difference in bacterial evenness (P>0.05) ; Beta diversity showed differences in microbial community structure between the two groups (P<0.05 ). Single factor microbial association network analysis showed that there was a high correlation between Firmicutes and Bacteroidetes in the pneumoconiosis and dust exposed groups and other species, showing a positive correlation; The correlation between Proteobacteria and other species is high, showing a negative correlation. Conclusion: The structure and relative abundance of bacteria in lower respiratory tract were different between patients with pneumoconiosis and dust exposure, and the diversity of bacteria in lower respiratory tract increased in patients with pneumoconiosis, which may be related to disease status.

Influence of antimicrobial peptide biofunctionalized TiO2 nanotubes on the biological behavior of human keratinocytes and its antibacterial effect / 中华口腔医学杂志

Objective: To fabricate TiO2 nanotube material functionalized by antimicrobial peptide LL-37, and to explore its effects on biological behaviors such as adhesion and migration of human keratinocytes (HaCaT) and its antibacterial properties. Methods: The TiO2 nanotube array (NT) was constructed on the surface of polished titanium (PT) by anodization, and the antimicrobial peptide LL-37 was loaded on the surface of TiO2 nanotube (LL-37/NT) by physical adsorption. Three samples were selected by simple random sampling in each group. Surface morphology, roughness, hydrophilicity and release characteristics of LL-37 of the samples were analyzed with a field emission scanning electron microscope, an atomic force microscope, a contact angle measuring device and a microplate absorbance reader. HaCaT cells were respectively cultured on the surface of three groups of titanium samples. Each group had 3 replicates. The morphology of cell was observed by field emission scanning electron microscope. The number of cell adhesion was observed by cellular immunofluorescence staining. Cell counting kit-8 (CCK-8) assay was used to detect cell proliferation. Wound scratch assay was used to observe the migration of HaCaT. The above experiments were used to evaluate the effect of each group on the biological behavior of HaCaT cells. To evaluate their antibacterial effects, Porphyromonas gingivalis (Pg) was respectively inoculated on the surface of three groups of titanium samples. Each group had 3 replicates. The morphology of bacteria was observed by field emission scanning electron microscope. Bacterial viability was determined by live/dead bacterial staining. Results: A uniform array of nanotubes could be seen on the surface of titanium samples in LL-37/NT group, and the top of the tube was covered with granular LL-37. Compared with PT group [the roughness was (2.30±0.18) nm, the contact angle was 71.8°±1.7°], the roughness [(20.40±3.10) and (19.10±4.11) nm] and hydrophilicity (the contact angles were 22.4°±3.1° and 25.3°±2.2°, respectively) of titanium samples increased in NT and LL-37/NT group (P<0.001). The results of in vitro release test showed that the release of antimicrobial peptide LL-37 was characterized by early sudden release (1-4 h) and long-term (1-7 d) slow release. With the immunofluorescence, more cell attachment was found on NT and LL-37/NT than that on PT at the first 0.5 and 2.0 h of culture (P<0.05). The results of CCK-8 showed that there was no significant difference in the proliferation of cells among groups at 1, 3 and 5 days after culture. Wound scratch assay showed that compared with PT and NT group, the cell moved fastest on the surface of titanium samples in LL-37/NT group at 24 h of culture [(96.4±4.9)%] (F=35.55, P<0.001). A monolayer cells could be formed and filled with the scratch in 24 h at LL-37/NT group. The results of bacterial test in vitro showed that compared with the PT group, the bacterial morphology in the NT and LL-37/NT groups was significantly wrinkled, and obvious bacterial rupture could be seen on the surface of titanium samples in LL-37/NT group. The results of bacteria staining showed that the green fluorescence intensity of titanium samples in LL-37/NT group was the lowest in all groups (F=66.54,P<0.001). Conclusions: LL-37/NT is beneficial to the adhesion and migration of HaCaT cells and has excellent antibacterial properties, this provides a new strategy for the optimal design of implant neck materials.

Variation and interaction mechanism between active components in Rheum officinale and rhizosphere soil microorganisms under drought stress / 中国中药杂志

To explore the changes and the reaction mechanisms between soil microecological environment and the content of secon-dary metabolites of plants under water deficit, this study carried out a pot experiment on the 3-leaf stage seedlings of Rheum officinale to analyze their response mechanism under different drought gradients(normal water supply, mild, moderate, and severe drought). The results indicated that the content of flavonoids, phenols, terpenoids, and alkaloids in the root of R. officinale varied greatly under drought stresses. Under mild drought stress, the content of substances mentioned above was comparatively high, and the content of rutin, emodin, gallic acid, and(+)-catechin hydrate in the root significantly increased. The content of rutin, emodin, and gallic acid under severe drought stress was significantly lower than that under normal water supply. The number of species, Shannon diversity index, richness index, and Simpson index of bacteria in the rhizosphere soil were significantly higher than those in blank soil, and the number of microbial species and richness index decreased significantly with the aggravation of drought stresses. In the context of water deficit, Cyanophyta, Firmicutes, Actinobacteria, Chloroflexi, Gemmatimonadetes, Streptomyces, and Actinomyces were the dominant bacteria in the rhizosphere of R. officinale. The relative content of rutin and emodin in the root of R. officinale was positively correlated with the relative abundance of Cyanophyta and Firmicutes, and the relative content of(+)-catechin hydrate and(-)-epicatechin gallate was positively correlated with the relative abundance of Bacteroidetes and Firmicutes. In conclusion, appropriate drought stress can increase the content of secondary metabolites of R. officinale from physiological induction and the increase in the association with beneficial microbe.

The structure and function analysis of bacterial community during aerobic composting of chicken manure / 生物工程学报

In order to determine the changes of bacterial community structure and function in the early, middle and late stage of aerobic composting of chicken manure, high-throughput sequencing and bioinformatics methods were used to determine and analyze the 16S rRNA sequence of samples at different stages of composting. Wayne analysis showed that most of the bacterial OTUs in the three composting stages were the same, and only about 10% of the operational taxonomic units (OTUs) showed stage specificity. The diversity indexes including Ace, Chao1 and Simpson showed a trend of increasing at first, followed by decreasing. However, there was no significant difference among different composting stages (P < 0.05). The dominant bacteria groups in three composting stages were analyzed at the phylum and genus levels. The dominant bacteria phyla at three composting stages were the same, but the abundances were different. LEfSe (line discriminant analysis (LDA) effect size) method was used to analyze the bacterial biological markers with statistical differences among three stages of composting. From the phylum to genus level, there were 49 markers with significant differences among different groups. The markers included 12 species, 13 genera, 12 families, 8 orders, 1 boundary, and 1 phylum. The most biomarkers were detected at early stage while the least biomarkers were detected at late stage. The microbial diversity was analyzed at the functional pathway level. The function diversity was the highest in the early stage of composting. Following the composting, the microbial function was enriched relatively while the diversity decreased. This study provides theoretical support and technical guidance for the regulation of livestock manure aerobic composting process.

Construction of "lock-key" biological living material based on double engineered bacteria and its application on intestinal retention in vivo / 生物工程学报

At present, the research of biological living materials mainly focuses on applications in vitro, such as using a single bacterial strain to produce biofilm and water plastics. However, due to the small volume of a single strain, it is easy to escape when used in vivo, resulting in poor retention. In order to solve this problem, this study used the surface display system (Neae) of Escherichia coli to display SpyTag and SpyCatcher on the surface of two strains, respectively, and constructed a double bacteria "lock-key" type biological living material production system. Through this force, the two strains are cross-linked in situ to form a grid-like aggregate, which can stay in the intestinal tract for a longer time. The in vitro experiment results showed that the two strains would deposit after mixing for several minutes. In addition, confocal imaging and microfluidic platform results further proved the adhesion effect of the dual bacteria system in the flow state. Finally, in order to verify the feasibility of the dual bacteria system in vivo, mice were orally administrated by bacteria A (p15A-Neae-SpyTag/sfGFP) and bacteria B (p15A-Neae-SpyCatcher/mCherry) for three consecutive days, and then intestinal tissues were collected for frozen section staining. The in vivo results showed that the two bacteria system could be more detained in the intestinal tract of mice compared with the non-combined strains, which laid a foundation for further application of biological living materials in vivo.

Advances in bacterial adsorption and transport of aromatic compounds / 生物工程学报

Aromatic compounds are a class of organic compounds with benzene ring(s). Aromatic compounds are hardly decomposed due to its stable structure and can be accumulated in the food cycle, posing a great threat to the ecological environment and human health. Bacteria have a strong catabolic ability to degrade various refractory organic contaminants (e.g., polycyclic aromatic hydrocarbons, PAHs). The adsorption and transportation are prerequisites for the catabolism of aromatic compounds by bacteria. While remarkable progress has been made in understanding the metabolism of aromatic compounds in bacterial degraders, the systems responsible for the uptake and transport of aromatic compounds are poorly understood. Here we summarize the effect of cell-surface hydrophobicity, biofilm formation, and bacterial chemotaxis on the bacterial adsorption of aromatic compounds. Besides, the effects of outer membrane transport systems (such as FadL family, TonB-dependent receptors, and OmpW family), and inner membrane transport systems (such as major facilitator superfamily (MFS) transporter and ATP-binding cassette (ABC) transporter) involved in the membrane transport of these compounds are summarized. Moreover, the mechanism of transmembrane transport is also discussed. This review may serve as a reference for the prevention and remediation of aromatic pollutants.

Application of cell-free transcription and translation system in CRISPR technologies and the associated biosensors / 生物工程学报

Cell-free transcription and translation (TXTL) system is a cell extract-based system for rapid in vitro protein expression. The system bypasses routine laboratory processes such as bacterial transformation, clonal screening and cell lysis, which allows more precise and convenient control of reaction substrates, reduces the impact of bacteria on protein production, and provides a high degree of versatility and flexibility. In recent years, TXTL has been widely used as an emerging platform in clusterd regularly interspaced short palindromic repeat (CRISPR) technologies, enabling more rapid and convenient characterization of CRISPR/Cas systems, including screening highly specific gRNAs as well as anti-CRISPR proteins. Furthermore, TXTL-based CRISPR biosensors combined with biological materials and gene circuits are able to detect pathogens through validation of related antibiotics and nucleic acid-based markers, respectively. The reagents can be freeze-dried to improve portability and achieve point-of-care testing with high sensitivity. In addition, combinations of the sensor with programmable circuit elements and other technologies provide a non-biological alternative to whole-cell biosensors, which can improve biosafety and accelerate its application for approval. Here, this review discusses the TXTL-based characterization of CRISPR and their applications in biosensors, to facilitate the development of TXTL-based CRISPR/Cas systems in biosensors.

Discovery, structure and function of plasmid mediated shufflon / 生物工程学报

Antimicrobial resistance has become a major public health issue of global concern. Conjugation is an important way for fast spreading drug-resistant plasmids, during which the type Ⅳ pili plays an important role. Type Ⅳ pili can adhere on the surfaces of host cell and other medium, facilitating formation of bacterial biofilms, bacterial aggregations and microcolonies, and is also a critical factor in liquid conjugation. PilV is an adhesin-type protein found on the tip of type Ⅳ pili encoded by plasmid R64, and can recognize the lipopolysaccharid (LPS) molecules that locate on bacterial membrane. The shufflon is a clustered inversion region that diversifies the PilV protein, which consequently affects the recipient recognition and conjugation frequency in liquid mating. The shufflon was firstly discovered on an IncI1 plasmid R64 and has been identified subsequently in plasmids IncI2, IncK and IncZ, as well as the pathogenicity island of Salmonella typhi. The shufflon consists of four segments including A, B, C, and D, and a specific recombination site named sfx. The shufflon is regulated by its downstream-located recombinase-encoding gene rci, and different rearrangements of the shufflon region in different plasmids were observed. Mobile colistin resistance gene mcr-1, which has attracted substantial attentions recently, is mainly located in IncI2 plasmid. The shufflon may be one of the contributors to fast spread of mcr-1. Herein, we reviewed the discovery, structure, function and prevalence of plasmid mediated shufflon, aiming to provide a theoretical basis on transmission mechanism and control strategy of drug-resistant plasmids.

Establishment and evaluation of a method for phages enrichment by ferric trichloride-polyvinylidene fluoride membrane filter / 中华预防医学杂志

Objective: To establish and evaluate a method of enriching bacteriophages in natural water based on ferric trichloride-polyvinylidene fluoride (FeCl3-PVDF)membrane filter. Methods: Based on the principle of flocculation concentration, the method of recovering bacteriophage from water sample was established by using iron ion flocculation combined with membrane filter. The titer of phage was determined by Agar double layer method. The recovery efficiency of phage was detected by phage fluorescence staining and real-time fluorescence PCR reaction. Water samples from different sources were collected for simulation experiment to evaluate the enrichment effect. At the same time, the sewage discharged from hospitals was taken as the actual water sample, and the common clinical drug-resistant bacteria were used as the host indicator bacteria to further analyze the enrichment effect of FeCl3-PVDF membrane filter rapid enrichment method on the bacteriophage in natural water samples. Results: The method of enrichment of bacteriophages in natural water by iron ion concentration 50 mg/L and PVDF membrane filter was established. The recovery rate of this method for bacteriophage was 93%-100%. Under the multi-functional microscope, it was found that the bacteriophage of the enriched water sample increased significantly and the fluorescence value of the enriched water sample determined by the enzyme labeling instrument was about 13 times as high as that before enrichment. After concentration of the actual water samples from the hospital drainage, the positive rate of bacteriophage isolation in the concentrated group and the non-concentrated group was 23% and 4%, and the fluorescence value in the concentrated group was 2-24 times as high as that of the non-concentrated group. Conclusion: The method of FeCl3-PVDF membrane filter is a simple, efficient and rapid method for enriching bacteriophages in different water samples.

Research progress of CRISPR/Cas biosensors based on different signal amplification strategies / 中华预防医学杂志

CRISPR/Cas(the clustered regularly interspaced short palindromic repeats-CRISPR associated)system exists in most bacteria and all archaea. It is an important strategy for bacteria and archaea to resist foreign nucleic acid invasion and use for self-defense. The CRISPR/Cas system is a simple, fast, and specific diagnostic tool, which is widely used in agriculture, industry, animal husbandry, and medicine. This article mainly introduces and discusses recently advantages and limitations of biosensors combining CRISPR/Cas system with fluorescence, visualization and surface enhanced raman related technologies, as well as future research directions.

Bacteriological analysis of nasal secretions in patients with nasal lymphoma / 临床耳鼻咽喉头颈外科杂志

Objective:To investigate the etiological characteristics of nasal bacterial infection in patients with nasal lymphoma. Methods:The results of bacterial culture of nasal secretions from 39 healthy people and 86 patients with nasal lymphoma in the Affiliated Hospital of Qingdao University from January 2019 to June 2022 were retrospectively analyzed, and the differences in nasal bacteria distribution between nasal lymphoma and healthy people were analyzed and compared. Results:Corynebacterium（38.90%） was the most common bacteria in the nasal cavity of healthy people, followed by coagulase-negative Staphylococcus（31.95%）, Staphylococcus epidermidis（15.28%） and Staphylococcus aureus（6.95%）. The most common bacteria in nasal lymphoma patients was Staphylococcus aureus（30.37%）, followed by Corynebacterium（9.63%）, Staphylococcus epidermidis（7.41%） and coagulase negative Staphylococcus（6.67%）. A total of 81 nasal lymphoma patients were detected with bacteria, positive rate is as high as 94.19%（81/86）. Conclusion:Staphylococcus aureus is the main pathogenic bacteria in nasal secretion of patients with nasal lymphoma, which provides guiding significance for the clinical prevention and treatment of nasal lymphoma complicated with infection or not.

Effects of rumen microorganisms on the decomposition of recycled straw residue / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Recently, returning straw to the fields has been proved as a direct and effective method to tackle soil nutrient loss and agricultural pollution. Meanwhile, the slow decomposition of straw may harm the growth of the next crop. This study aimed to determine the effects of rumen microorganisms (RMs) on straw decomposition, bacterial microbial community structure, soil properties, and soil enzyme activity. The results showed that RMs significantly enhanced the degradation rate of straw in the soil, reaching 39.52%, which was 41.37% higher than that of the control on the 30th day after straw return. After 30 d, straw degradation showed a significant slower trend in both the control and the experimental groups. According to the soil physicochemical parameters, the application of rumen fluid expedited soil matter transformation and nutrient buildup, and increased the urease, sucrase, and cellulase activity by 10%‒20%. The qualitative analysis of straw showed that the hydroxyl functional group structure of cellulose in straw was greatly damaged after the application of rumen fluid. The analysis of soil microbial community structure revealed that the addition of rumen fluid led to the proliferation of Actinobacteria with strong cellulose degradation ability, which was the main reason for the accelerated straw decomposition. Our study highlights that returning rice straw to the fields with rumen fluid inoculation can be used as an effective measure to enhance the biological value of recycled rice straw, proposing a viable solution to the problem of sluggish straw decomposition.