A leptospirose é uma zoonose causada por bactérias patogênicas do gênero Leptospira. Apesar de sua importância, a patogênese e virulência permanecem não elucidadas. As leptospiras não apresentam proteases conhecidas de degradação de matriz extracelular, atividade crucial para a penetração e disseminação nos hospedeiros. Assim, foi proposta a investigação da interação de leptospiras com plasminogênio/plasmina e as implicações para a infecção. As leptospiras capturam plasminogênio na superfície, e este é convertido à plasmina por ativadores do hospedeiro. A plasmina associada propicia degradação de componentes de matriz extracelular, habilidade de penetração e evasão imune. Adicionalmente, as leptospiras estimulam a expressão de ativadores de plasminogênio e metaloproteases de matriz. Os resultados contribuem para o conhecimento do processo infeccioso das leptospiras, descrevendo um novo mecanismo de patogenicidade. / Leptospirosis is a zoonosis caused by pathogenic bacteria from genus Leptospira. Despite its importance, the pathogenicity and virulence remain to be elucidated. The leptospires do not present known proteases able to degrade extracellular matrix, an activity essential for the penetration and dissemination within the hosts. Therefore, we proposed the investigation of the leptospiral interaction with plasminogen/plasmin and its implications for infection. Leptospires capture plasminogen on the surface, which is converted to plasmin by hosts activators. Surface-bound plasmin confers extracellular matrix components degradation, penetration ability and immune evasion. Additionally, leptospires stimulate plasminogen activators and matrix metaloproteases expressions. The results constitute one possible mechanism that contributes to the invasion process and the rapid dissemination of Leptospira.

Thesis (M.Sc.(Microbiology)) -- University of Limpopo, 2019 / Salmonella enterica serovar Gallinarum biovars Gallinarum, and Pullorum, Pasteurella multocida and Escherichia coli are among the most important pathogens in poultry and are the causal agents of fowl typhoid, pullorum disease, fowl cholera and collibacillosis in poultry. The present study was designed to identify and determine the distribution of these pathogens in household-raised chickens and their antibiotic and virulence profiles. For this purpose, 40 chickens were bought from household families at Ga-Dikgale, GaMolepo and Ga-Mphahlele in the Capricorn district of Limpopo Province and sacrificed for sampling. Tissues including breast meat, lungs, small and large intestines were harvested from each chicken. Bacteria associated with these samples were cultured in selective bacteriological media followed by biotyping using matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) for identification. Out of a total of 160 tissue samples evaluated, E. coli and Salmonella were detected in these tissues. Furthermore, determination of the pathogenic E. coli and Salmonella strains at species level using primer sets that target selected genes of interest in the polymerase chain reaction (PCR) assay was employed. The invA gene, a confirmatory gene for Salmonella species was detected in all the Salmonella isolates using PCR. For the pathogenic E. coli, astA, eae, hlyA, fIiCH7, stxI and the fimbrial genes (F6 and F41) were detected in some of the E. coli isolates recovered from the samples. Disk diffusion test was also performed to determine the antibiotic susceptibility of the bacteria. The results from the current samples reveals that there is a high distribution of Salmonella and pathogenic E. coli in these areas and therefore further epidemiological and identification studies are needed to determine these organisms at species level and investigate their pathogenicity. The antimicrobial susceptibly data generated from this study can be a valuable reference to veterinarians for treating bacterial diseases in poultry.

Yeast probiotics and paraprobiotics, live and inactivated yeast cells, respectively, improve health and performance of livestock by stabilizing the intestinal microbial community. They have also been used for infection prevention and treatment. Despite much research already conducted, the mechanism of direct antagonism, or adhesion of bacteria to the probiotic/paraprobiotic, is under characterized. Additionally, it is unknown which probiotic/paraprobiotic is optimal to use for specific infections. The interactions between the yeast and certain pathogens were analyzed qualitatively with scanning electron microscopy (SEM) and quantitatively with membrane filtration assays. Gram-positive bacteria were found to exhibit specificity under SEM. Through membrane filtration, Listeria monocytogenes exhibited binding to all samples (P<0.05), while Salmonella Typhimurium exhibited binding (P<0.001) with all samples except with 2338. Escherichia coli O157:H7 only bound to the probiotics (P<0.001). With a better understanding of how specific yeast probiotics and paraprobiotics interact with bacteria, specific therapies can be administered to combat infections.

Several antibiotics that prevent translation by binding to ribosomal subunits have been shown to also inhibit ribosomal subunit assembly (Champney and Tober 2003). The aminoglycoside hygromycin B was examined in Escherichia coli cells for inhibitory effects on translation and ribosomal subunit assembly. The streptogramin antibiotics quinupristin-dalfopristin and XRP 2868 (NXL 103) were examined for similar effects on these 2 cellular functions in antibiotic-resistant strains of Haemophilus influenzae, Staphylococcus aureus, and Streptococcus pneumoniae. Pulse chase experiments were performed which verified slower rates of ribosomal subunit formation in drug treated cells. Hygromycin B exhibited a concentration dependent inhibitory effect on viable cell number, growth rate, protein synthesis and 30S and 50S subunit formation. 16S rRNA specific probes hybridized to rRNA fragments in cells treated with hygromycin B. RNase II and RNase III deficient strains of E. coli exhibited the most accumulation of 16S rRNA fragments upon treatment with hygromycin B. Examination of total RNA from treated cells showed an increase in RNA corresponding to precursor to the 16S rRNA while 16S rRNA decreased. There was also an increase in small fragment RNA. Hygromycin B was a more effective inhibitor of translation than ribosomal subunit formation in E. coli. Two streptogramin antibiotics were compared for inhibitory effects in antibiotic-resistant Haemophilus influenzae, Staphylococcus aureus, and Streptococcus pneumoniae. IC50 values for XRP 2868 were several fold lower than those of quinupristin-dalfopristin for inhibition of cell viability, protein synthesis, and ribosomal subunit formation. Both antibiotics revealed a concentration dependent inhibitory effect on cellular functions including 50S ribosomal subunit formation in the three organisms examined. XRP 2868 inhibited both 50S ribosomal subunit assembly and translation. XRP 2868 was effective against MRSA and was a better inhibitor in each of the antibiotic resistant strains examined compared with quinupristin-dalfopristin.

This study is entitled, “determination of the presence of antibiotics and bacterial pathogens with their susceptibility in fresh milk from the San Nicolas community and the cities of Penipe, Guano, and Chambo in the Chimborazo province”. It was funded by the Benson Institute of Brigham Young University and Nestle. The objectives were: 1) to determine the presence of antibiotics in fresh milk from the San Nicolas community and the cities of Penipe, Guano, and Chambo, 2) to determine the presence of bacterial pathogens in milk from the places previously mentioned, and 3) to detect the susceptibility of the isolated bacteria to the most commonly used antibiotics on the market. With regard to the presence of antibiotics, they were only found once in Chambo (beta-lactams) and once in San Nicolas (tetracycline). Three types of pathogenic bacteria were found: Escherichia coli, Streptococcus agalactiae, and Staphylococcus aureus. Only one bacterium was found that has not been studied: Pseudomonas aureoqinosa. This bacterium was found in milk from San Nicolas. With regard to the susceptibility of the bacteria to antibiotics, it was found that Escherichia coli was sensitive to the following antibiotics: nitrofurans, erimethoprina, trimethoprim and sulfonamides, and tetracyclines and spectomicina. Escherichia coli also showed medium sensitivity to chloramphenicol as well as resistance to ampicillin and sulfonamides. Streptococcus agalactiae was sensitive to lincomycin, neomycin, enrofloxacin, and clindamycin. It also showed medium sensitivity to erythromycin and low tetracycline and penicillin as well as resistance to gentamicin. Lastly, Staphylococcus aureus was sensitive to neomycin and enrofloxacin. In addition, it showed medium sensitivity to lincomycin, gentamicin, clindamycin, and tetracycline as well as resistance to low erythromycin and penicillin. This study was done over a four month period with at least 12 samples being collected from each area. A total of 60 samples were analyzed. Based on the results obtained, our recommendation is that hygienic measures are taken in obtaining and transporting milk to eliminate the growth of bacterial pathogens. Also, we recommend the use of the effective antibiotics listed above in order to completely eradicate these microorganisms.

京都大学 / 新制・課程博士 / 博士(工学) / 甲第24898号 / 工博第5178号 / 新制||工||1988(附属図書館) / 京都大学大学院工学研究科都市環境工学専攻 / (主査)教授 伊藤 禎彦, 教授 松田 知成, 教授 越後 信哉 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DGAM

Recent demands for minimally-processed foods, has led to the exploration of plant-derived essential oil (EO) compounds as an alternative means of preservation. While some of these compounds are effective against foodborne pathogens, their strong aroma and "spicy" flavor are not compatible with the flavor of juice. The purpose of this research was to evaluate the antimicrobial activity of three EO compounds (thymol, eugenol, and trans-cinnamaldehyde) alone and in combination with three naturally-occurring apple aroma compounds (hexanal, trans-2-hexenal and 1-hexanol) in order to identify combinations that lower the concentrations needed to destroy foodborne pathogens in apple juice. The standard agar dilution method (SAD) and the Spiral Gradient Endpoint method (SGE) were compared for their abilities to determine minimum inhibitory concentrations (MIC) of the EO compounds. Both methods produced similar patterns of inhibition; however, the MICs produced by the SGE system were significantly higher than those produced by the SAD method of analysis (P<0.05). Since the results produced by the SAD method were more comparable with those published in literature, this method was selected for further testing. In general, the EO compounds were significantly more effective against the test pathogens (Listeria monocytogenes, Salmonella Typhimurium and Staphylococcus aurues) than were the apple aroma compounds (P<0.05). Cinnamaldehye exhibited the highest degree of activity, followed by thymol and eugenol. Eugenol was the only compound that acted synergistically with the apple aroma compounds. The most effective compounds (cinnamaldehyde, eugenol and trans-2-hexenal) were then used to inactivate L. monocytogens and S. Typhimurium in preservative-free apple juice. In most cases, treatment with 0.05% of each compound resulted in a 5 log CFU/ml reduction in bacterial numbers following one day of storage at 4°C or 25°C. Likewise, treatment with antimicrobial combinations (containing 0.025% of trans-2-hexenal in combination with 0.025% trans-cinnamaldehyde or eugenol) also resulted in a 5 log CFU/ml reduction in bacterial numbers, following one day of storage at 4°C or 25°C. Since these combinations contained half the effective concentration of the essential oil compounds, they may be used to preserve the microbial quality of apple juice, while reducing the likelihood of off flavors in the final juice product. / Ph. D.

Hot peppers are gaining popularity in the United States as both a vegetable and a spice. In 2008, jalapeño peppers were involved in a multistate outbreak of Salmonella Saintpaul. This is the first outbreak implicating jalapeño as a vehicle for foodborne illness. Hot peppers contain many compounds thought to possess antimicrobial characteristics. This research was conducted to provide more information on the interactions of pathogenic bacteria and jalapeño peppers, as well as to identify properties of Capsicum annuum that affect bacterial survival, growth, and inhibition. Behavior of pathogens associated with jalapeños was investigated by inoculating jalapeño fruits with a cocktail of Listeria monocytogenes, Salmonella enterica, or Escherichia coli O157:H7 on the intact external surface, injured external surface, or intact internal cavity and storing the jalapeños at 7°C or 12°C. Intact external jalapeñosurfaces did not support the growth of the bacteria tested under storage conditions of 7°C. However, L. monocytogenes populations remained detectable throughout the 2 week study. At 7°C, pathogenic bacteria were able to survive but not grow on injured and internally inoculated jalapeño, but populations increased at 12°C (p=0.05). The most supportive growth environment for the pathogenic bacteria was the internal cavity of jalapeño held at 12°C. This study demonstrated the importance of intact uninjured produce and proper storage temperatures for food microbial safety. Inhibitory properties of jalapeños were studied by making extracts from fresh jalapeño peppers to test for antimicrobial activity. A disk diffusion assay determined that the extracts were capable of inhibiting the growth of the pathogenic bacteria tested. Listeria monocytogenes was especially sensitive to the extracts. jalapeño extracts were fractionated using HPLC and used for inhibition assays using disk diffusion and growth curve generation. Two fractions stimulated bacterial growth (p=0.05), while two other fractions inhibited bacterial growth. The inhibitory fractions were separated further using HPLC and tested for antimicrobial activity. Fraction E1 suppressed the growth of L. monocytogenes. HPLC-MS analysis revealed that Fraction E1 contained compounds known as capsianosides. To prove that inhibition is caused by capsianoside(s) and determine minimum inhibitory concentrations, a method to isolate the pure compound should be developed. / Ph. D.

Thesis (M. Tech.(Biomedical Technology)) - Central University of technology, Free State, 2011 / The existence of geophagia from as early as 460 BC up to now, makes it relevant to investigate all aspects related to geophagia. Geophagia is a direct route for potential transmission of pathogens to the human host, through the ingestion of soil. Soil-borne diseases in humans are causing growing concern as sewage disposal, which involve sewage sludge and waste water drainage from these plants, is on the increase. It is estimated that approximately seven million tons of sewage sludge is produced annually and that 54% of this sewage sludge is introduced into soil. Data on enteric infection in humans caused by contamination from soil is limited and need further investigation. The aim of the study was, therefore, to collect information on the microbiological presence in geophagic soil in the Qwa-Qwa district. Objectives included the collecting of information regarding various sampling sites in the Qwa-Qwa district and also soil samples sold by vendors, investigation of the prevalence of known human pathogenic bacteria and fungi in geophagic soil, investigating the culturability of Salmonella enteritidis in geophagic soil in comparison with the viability of these organisms in soil for long periods of time, investigating potential antimicrobial activity of geophagic soil, as some of the geophagists are convinced that the geophagic soils have medicinal properties, and to determine the microbial diversity of geophagic soils, which can not be accomplished by conventional microbial culturing methods.

Thesis (MSc (Microbiology))--Stellenbosch University, 2008. / Mastitis is considered to be the most costly disease affecting the dairy industry. Management strategies involve the extensive use of antibiotics to treat and prevent this disease. Prophylactic dosages of antibiotics used in mastitis control programmes could select for strains with resistance to antibiotics. In addition, a strong drive towards reducing antibiotic residues in animal food products has lead to research in finding alternative antimicrobial agents. Streptococcus macedonicus ST91KM, isolated from bulgarian goat yoghurt, produces the bacteriocin macedocin ST91KM with a narrow spectrum of activity against Grampositive bacteria. These include mastitis pathogens Streptococcus agalactiae, Streptococcus dysgalactiae, Streptococcus uberis, Staphylococcus aureus and Staphylococcus epidermidis as well as Lactobacillus sakei and Micrococcus varians. Macedocin ST91KM is, according to tricine-SDS PAGE, between 2.0 and 2.5 kDa in size. The activity of macedocin ST91KM remained unchanged after 2 h of incubation at pH 2.0 to 10.0 and 100 min at 100 °C. The peptide was inactivated after 20 min at 121 °C and when treated with pronase, pepsin and trypsin. Treatment with α-amylase had no effect on activity, suggesting that the mode of action does not depend on glycosylation. Precipitation with 60 % saturated ammonium sulphate, followed by Sep-Pak C18 separation recovered 43 % of macedocin ST91KM. Amplification of the genome of strain ST91KM with primers designed from the sequence of the macedocin prescursor gene (mcdA) produced two fragments (approximately 375 and 220 bp) instead of one fragment of 150 bp recorded for macedocin produced by S. macedonicus ACA-DC 198. Strain ACA-DC 198 was not available. However, the DNA fragment amplified from strain LMG 18488 (ACA-DC 206), genetically closely related to strain ACADC 198, revealed 99 % homology to the mcdA of S. macedonicus ACA-DC 198 (accession number DQ835394). Macedocin ST91KM may thus be a related bacteriocin described for S. macedonicus. The peptide adsorbed equally well (66 %) to L. sakei LMG13558 and insensitive cells, e.g. Enterococcus faecalis BFE 1071 and FAIR E92, and Streptococcus caprinus ATCC 700066. Optimal adsorption of macedocin ST91KM was recorded at 37 °C and 45 °C and at pH of 8 - 10. Addition of solvents decreased adsorption by 50%, suggesting that the receptors to which the bacteriocin binds have lipid moieties. The addition of MgCl2, KI and Na2CO3 completely prevented adsorption of macedocin ST91KM to the target cells, possibly due to competitive ion adsorption on the bacterial cell surface. The peptide has a bacteriocidal mode of action, resulting in lysis and the release of DNA and β-galactosidase. Atomic force microscopy of sensitive cells treated with macedocin ST91KM have shown deformation of the cell structure and developing of irregular surface areas. Antimicrobial susceptibility patterns were evaluated against eighteen mastitis pathogens. All isolates tested were resistant to methicillin and oxacillin, but had minimum inhibitory concentrations (MICs) falling in the intermediate and susceptible range against erythromycin. S. agalactiae and S. epidermidis had the highest sensitivity to macedocin ST91KM. A teat seal preparation containing macedocin ST91KM effectively released bacteriocin inhibiting the growth of the bacterial pathogen. Macedocin ST91KM could form the basis for an alternative dry cow therapy to prevent mastitis infections in dairy cows, as it is effective against pathogens that display resistance to conventional antibiotic therapy.