DOCSLIB.ORG

Streptococcus Laboratory General Methods

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text

Load more

The reference used for compiling the methods in Section I is: Murray, P.R., Baron, E. J., Jorgensen, J.J., Pfaller, M.A., and Yolken, R.H. Manual of Clinical Microbiology, 8th ed. ASM Press: Washington, DC, 2003. The Streptococcus species identification methods in Section II were compiled by Dr. Lynn Shewmaker. Also thanks to input from several individuals, including Richard Facklam and Lucia Teixeira. Section I. 1. Accuprobe-Enterococcus Test………….………..4 2. Accuprobe-Pneumococcus Test …………..….….4 3. Acid formation in carbohydrate broth..................5 4. Arginine Hydrolysis……………………….……….6 5. Bacitracin Test……………………………………..7 6. Bile-esculin Test…………………………………...8 7. Bile solubility Test …………………………………9 8. CAMP Test…......................................................10 9. Catalase Test......................................................11 10. Clindamycin test………………………………….12 11. Esculin hydrolysis……………………………….. 13 12. Gas from MRS broth……………………………...14 13. Gram Stain………………………………………...15 14. Growth at 10 & 45C……………………………. 17 15. Hemolysis………………………………………….18 16. Hippurate hydrolysis…………………………… 19 17. Lancefield Group Antigen………………………..20 18. Leucine amino peptidase (LAP)…………………21 19. Litmus Milk Test…………………………………..22 20. Motility………………………………………………23 21. 6.5% NaCl Tolerace Test...................................24 22. Optochin…………………………………………….25 23. Pigmentation....................................................... 26 24. Pyridoxal Requirement Test (Vitamin B6)……….27 25. Pyrrolidonlarylamindase (PYR)............................28 26. Pyruvate Utilization Test.......................................29 27. Satellite Test…………..........................................30 28. Starch hydrolysis..................................................31 29. Sucrose agar (extracellular polysaccharides)….. 32 30. Sucrose broth (5%).............................................. 33 31. Tellurite tolerance.................................................34 32. Urea hydrolysis.....................................................35 33. Vancomycin test................................................... 36 34. Voges-Proskauer.................................................. 37 1 Centers for Disease Control and Prevention Updated March, 2014 Section II. Identification Procedures for Catalase-Negative Gram-positive Cocci, page 54. A. General Discription of the genera identified in this lab, page 54. Table 1. Identification of gram-positive genera...............................43 Beta-hemolytic streptococci............................................................48 Table 2. Identification and reporting of β-hemolytic streptococci....50 Table 3. Identification of atypical group A strep..............................51 Table 4. Identification of the S. anginous group..............................52 Non-beta-hemolytic streptococci.....................................................53 Streptococcus pneumoniae.............................................................53 Table 5. Identification non-β-hemolytic strep...................................54 Table 6. Characteristics of viridans streptococci.............................55 Table 7. Identification of nutritionally variant strep...........................56 B. Identification of Enterococcus and Vagococcus……………………………57 Table 8. Identification of Enterococcus and Vagococcus species…58 C. Identification of Lactococcus species..................................................61 Table 9. Identification of Lactococcus species.................................61 D. Identification of Gemella species..........................................................62 Table 10. Identification of Gemella species.....................................62 E. Identification of Pediococcus and Tetragenococcus species............62 Table 11. Identification of Pediococcus and Tetragenococcus.........63 F. Identification of Leuconostoc species..................................................64 Table 12. Identification of Leuconostoc species……………………..64 G. Identification of Aerococcus and Helococcus.....................................65 Table 13. Identification of Aerococcus and Helococcus…………….65 H. Identification of Globicatella..................................................................66 Table 14. Identification of Globicatella..............................................67 I. Identification of Dolosigranulum, Ignavigranum,Facklamia, and Alloicococcus Table 15. Identification of Dolosigranulum, Ignavigranum, Facklamia, and Alloicococcus..........................................................68 Table 16. Rapid ID-32 for unusual gram positive bacteria................69 2 OUTLINE OF THE WORK FLOW FOR THE STREPTOCOCCUS LABORATORY 1. If the culture is an unidentified gram-positive coccus, an Enterococcus, viridans Streptococcus, or of unknown identity (basically includes all cultures other than pneumococci, ß-hemolytic streptococci, and nutritionally variant streptococci), inoculate the following media. Inoculate a trypticase soy 5% sheep blood agar plate by streaking a heavy inoculum onto one-fourth of the plate and streak the remaining portion for isolated colonies. Place a vancomycin disk on the heaviest part of the inoculum, and put the plate into a candle extinction jar or a CO2 incubator for 18 to 24 h at 35C. 2. If the culture is identified as a beta-hemolytic streptococcus, or group A, B, C, F, or G streptococci, inoculate a trypticase soy 5% sheep blood agar plate and place a bacitracin disk on the heavy part of the growth. All plates should be incubated in a candle extinction jar or CO2 incubator for 18 h at 35ΕC. For most cultures submitted as group A streptococci, those that appear pure on the submitted culture, a 30 ml and a 5 ml Todd-Hewitt broth (THB) should be inoculated. Place the 5 ml THB in a 30Ε C incubator or leave at room temperature for 1-3 days. The 5 ml THB is used for T-agglutination typing of group A streptococci. Place the 30 ml broth at 35ΕC for 16 to 18 h. Do not incubate longer than 18 h. For beta-hemolytic streptococci other than group A, a 30 ml THB can also be inoculated and placed at 35ΕC for 18 to 24 h. Some strains may take more than one day of incubation, no harm is done by incubating these broths longer than 24 to 72 h. The 30 ml THB is used for serogrouping and serotyping in some cases. 4. If the culture is submitted as a nutritionally deficient Streptococcus (NVS), inoculate an entire trypticase soy blood agar plate with the culture. Perpendicular to this streak, carefully make a single streak with a Staphylococcus aureus culture. This test will determine if the unknown culture forms satellite colonies adjacent to the staphylococci, a characteristic of all NVS. Incubate the plate in CO2 or a candle extinction jar at 35Ε C for 24 to 48 h. Examples of unusual or unexpected results: The blood agar plates used in the manner described above are for checking for purity of cultures. If any of the results are unusual or not expected or the culture is contaminated the test must be repeated. Vancomycin resistant streptococci or other unknowns other than leuconostocs and pediococci, which are intrinsically vancomycin resistant. 3 AccuProbe-Enterococcus Test I. Principle The AccuProbe-Enterococcus test is used to aid in the identification of atypical enterococci and to help differentiate between Enterococcus and Lactococcus strains. II. Inoculum An overnight culture grown on blood agar incubated 35˚C in CO2. III. Reagents and Materials Genprobe Accuprobe Enterococcus Culture Identification Test, GEN-PROBE Inc. San Diego, CA IV. Procedure The test is performed according to the package insert instructions. V. Reading and Interpretaion Automated VI. Limitations Use care with the amount of colonies used. Too many colonies will result in a false positive test. VII. Quality Control Quality controls, positive and negative reactions are determined each day the test is determined. E. faecalis SS1273 and S. sanguinis SS910 are used as the positive and negative controls respecitively. AccuProbe-Pneumococcus Test I. Principle The AccuProbe-Pneumococcus test is used to aid in the identification of atypical pneumococci and to help differentiate between viridans Streptococcus strains. II. Inoculum An overnight culture grown on blood agar incubated 35˚C in CO2. III. Reagents and Materials Genprobe Accuprobe Pneumococcus Culture Identification Test, GEN-PROBE Inc. San Diego, CA IV. Procedure The test is performed according to the package insert instructions. V. Reading and Interpretaion Automated VI. Limitations Use care with the amount of colonies used. Too many colonies will result in a false positive test. VII. Quality Control Quality controls, positive and negative reactions are determined each day the test is determined. S. pneumonia and S. sanguinis SS910 are used as the positive and negative controls respecitively. 4 Acid Formation in Carbohydrate Broths I.Principle The ability of bacteria to form acid in some carbohydrate broths and not in others can be used in identification schemes. If the bacteria acidify the carbohydrate, the pH will change and the indicator (brom cresol purple) will turn yellow. II. Inoculum An overnight culture in Todd Hewitt broth incubated over night at 35˚ C or a fresh bacterial suspension in Todd Hewitt broth may be used as the inoculum. III. Reagents and Materials 1. Heart Infusion broth with 1% carbohydrate and 0.16 brom cresol purple indicator. Most of the carbohydrate broths are commercially available (Remel). An asterisk indicates those that are made by
Recommended publications
  • Probiotic Properties of Enterococcus Faecium CE5-1 Producing A

    Probiotic Properties of Enterococcus Faecium CE5-1 Producing A

    Czech J. Anim. Sci., 57, 2012 (11): 529–539 Original Paper Probiotic properties of Enterococcus faecium CE5-1 producing a bacteriocin-like substance and its antagonistic effect against antibiotic-resistant enterococci in vitro K. Saelim1, N. Sohsomboon1, S. Kaewsuwan2, S. Maneerat1 1Department of Industrial Biotechnology, Faculty of Agro-Industry, Prince of Songkla University, Hat Yai, Thailand 2Department of Pharmacognosy and Pharmaceutical Botany, Faculty of Pharmaceutical Sciences, Prince of Songkla University, Hat Yai, Thailand ABSTRACT: A bacteriocin-like substance (BLS) producing Enterococcus faecium CE5-1 was isolated from the gastrointestinal tract (GIT) of Thai indigenous chickens. Investigations of its probiotic potential were carried out. The competition between the BLS probiotic strain and antibiotic-resistant enterococci was also studied. Ent. faecium CE5-1 exhibited a good tolerance to pH 3.0 after 2 h and in 7% fresh chicken bile after 6 h, but the viability of Ent. faecium CE5-1 decreased by about 2–3 log CFU/ml after 2 h incubation in pH 2.5. It was susceptible to the antibiotics tested (tetracycline, erythromycin, penicillin G, and vancomycin). The maximum BLS production from Ent. faecium CE5-1 was observed at 15 h of cultivation. It showed activity against Listeria monocytogenes DMST17303, Pediococcus pentosaceus 3CE27, Lactobacillus sakei subsp. sakei JCM1157, and antibiotic-resistant enterococci. The detection by polymerase chain reaction (PCR) in the enterocin structural gene determined the presence of enterocin A gene in Ent. faecium CE5-1 only. Ent. faecium CE5-1 showed the highest inhibitory activity against two antibiotic-resistant Ent. faecalis VanB (from 6.68 to 4.29 log CFU/ml) and Ent.
  • Tryptose Blood Agar Base

    Tryptose Blood Agar Base

    Tryptose Blood Agar Base Intended Use Principles of the Procedure Tryptose Blood Agar Base is used with blood in isolating, Tryptose is the source of nitrogen, carbon and amino acids in cultivating and determining the hemolytic reactions of fastidi- Tryptose Blood Agar Base. Beef extract provides additional ous microorganisms. nitrogen. Sodium chloride maintains osmotic balance. Agar is the solidifying agent. Summary and Explanation Investigations of the nutritive properties of tryptose demon- Supplementation with 5-10% blood provides additional growth strated that culture media prepared with this peptone were factors for fastidious microorganisms and is used to determine superior to the meat infusion peptone media previously used hemolytic patterns of bacteria. for the cultivation of Brucella, streptococci, pneumococci, me- Formula ningococci and other fastidious bacteria. Casman1,2 reported Difco™ Tryptose Blood Agar Base that a medium consisting of 2% tryptose, 0.3% beef extract, Approximate Formula* Per Liter 0.5% NaCl, 1.5% agar and 0.03% dextrose equaled fresh beef Tryptose .................................................................... 10.0 g infusion base with respect to growth of organisms. The small Beef Extract ................................................................. 3.0 g amount of carbohydrate was noted to interfere with hemolytic Sodium Chloride ......................................................... 5.0 g Agar ......................................................................... 15.0 g reactions, unless the medium was incubated in an atmosphere *Adjusted and/or supplemented as required to meet performance criteria. of carbon dioxide. Tryptose Blood Agar Base is a nutritious infusion-free basal Directions for Preparation from medium typically supplemented with 5-10% sheep, rabbit or Dehydrated Product horse blood for use in isolating, cultivating and determining 1. Suspend 33 g of the powder in 1 L of purified water.
  • Detection of ESKAPE Pathogens and Clostridioides Difficile in Simulated

    Detection of ESKAPE Pathogens and Clostridioides Difficile in Simulated

    bioRxiv preprint doi: https://doi.org/10.1101/2021.03.04.433847; this version posted March 4, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. 1 Detection of ESKAPE pathogens and Clostridioides difficile in 2 Simulated Skin Transmission Events with Metagenomic and 3 Metatranscriptomic Sequencing 4 5 Krista L. Ternusa#, Nicolette C. Keplingera, Anthony D. Kappella, Gene D. Godboldb, Veena 6 Palsikara, Carlos A. Acevedoa, Katharina L. Webera, Danielle S. LeSassiera, Kathleen Q. 7 Schultea, Nicole M. Westfalla, and F. Curtis Hewitta 8 9 aSignature Science, LLC, 8329 North Mopac Expressway, Austin, Texas, USA 10 bSignature Science, LLC, 1670 Discovery Drive, Charlottesville, VA, USA 11 12 #Address correspondence to Krista L. Ternus, [email protected] 13 14 1 bioRxiv preprint doi: https://doi.org/10.1101/2021.03.04.433847; this version posted March 4, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. 15 1 Abstract 16 Background: Antimicrobial resistance is a significant global threat, posing major public health 17 risks and economic costs to healthcare systems. Bacterial cultures are typically used to diagnose 18 healthcare-acquired infections (HAI); however, culture-dependent methods provide limited 19 presence/absence information and are not applicable to all pathogens.
  • Current Trends of Enterococci in Dairy Products: a Comprehensive Review of Their Multiple Roles

    Current Trends of Enterococci in Dairy Products: a Comprehensive Review of Their Multiple Roles

    foods Review Current Trends of Enterococci in Dairy Products: A Comprehensive Review of Their Multiple Roles Maria de Lurdes Enes Dapkevicius 1,2,* , Bruna Sgardioli 1,2 , Sandra P. A. Câmara 1,2, Patrícia Poeta 3,4 and Francisco Xavier Malcata 5,6,* 1 Faculty of Agricultural and Environmental Sciences, University of the Azores, 9700-042 Angra do Heroísmo, Portugal; [email protected] (B.S.); [email protected] (S.P.A.C.) 2 Institute of Agricultural and Environmental Research and Technology (IITAA), University of the Azores, 9700-042 Angra do Heroísmo, Portugal 3 Microbiology and Antibiotic Resistance Team (MicroART), Department of Veterinary Sciences, University of Trás-os-Montes and Alto Douro (UTAD), 5001-801 Vila Real, Portugal; [email protected] 4 Associated Laboratory for Green Chemistry (LAQV-REQUIMTE), University NOVA of Lisboa, 2829-516 Lisboa, Portugal 5 LEPABE—Laboratory for Process Engineering, Environment, Biotechnology and Energy, Faculty of Engineering, University of Porto, 420-465 Porto, Portugal 6 FEUP—Faculty of Engineering, University of Porto, 4200-465 Porto, Portugal * Correspondence: [email protected] (M.d.L.E.D.); [email protected] (F.X.M.) Abstract: As a genus that has evolved for resistance against adverse environmental factors and that readily exchanges genetic elements, enterococci are well adapted to the cheese environment and may reach high numbers in artisanal cheeses. Their metabolites impact cheese flavor, texture, Citation: Dapkevicius, M.d.L.E.; and rheological properties, thus contributing to the development of its typical sensorial properties. Sgardioli, B.; Câmara, S.P.A.; Poeta, P.; Due to their antimicrobial activity, enterococci modulate the cheese microbiota, stimulate autoly- Malcata, F.X.
  • Fiber-Associated Spirochetes Are Major Agents of Hemicellulose Degradation in the Hindgut of Wood-Feeding Higher Termites

    Fiber-Associated Spirochetes Are Major Agents of Hemicellulose Degradation in the Hindgut of Wood-Feeding Higher Termites

    Fiber-associated spirochetes are major agents of hemicellulose degradation in the hindgut of wood-feeding higher termites Gaku Tokudaa,b,1, Aram Mikaelyanc,d, Chiho Fukuia, Yu Matsuuraa, Hirofumi Watanabee, Masahiro Fujishimaf, and Andreas Brunec aTropical Biosphere Research Center, Center of Molecular Biosciences, University of the Ryukyus, Nishihara, 903-0213 Okinawa, Japan; bGraduate School of Engineering and Science, University of the Ryukyus, Nishihara, 903-0213 Okinawa, Japan; cResearch Group Insect Gut Microbiology and Symbiosis, Max Planck Institute for Terrestrial Microbiology, 35043 Marburg, Germany; dDepartment of Entomology and Plant Pathology, North Carolina State University, Raleigh, NC 27607; eBiomolecular Mimetics Research Unit, Institute of Agrobiological Sciences, National Agriculture and Food Research Organization, Tsukuba, 305-8634 Ibaraki, Japan; and fDepartment of Sciences, Graduate School of Sciences and Technology for Innovation, Yamaguchi University, Yoshida 1677-1, 753-8512 Yamaguchi, Japan Edited by Nancy A. Moran, University of Texas at Austin, Austin, TX, and approved November 5, 2018 (received for review June 25, 2018) Symbiotic digestion of lignocellulose in wood-feeding higher digestion in the hindgut of higher termites must be attributed to termites (family Termitidae) is a two-step process that involves their entirely prokaryotic microbial community (5). endogenous host cellulases secreted in the midgut and a dense The gut microbiota of higher termites comprises more than bacterial community in the hindgut compartment. The genomes of 1,000 bacterial phylotypes, which are organized into distinc- the bacterial gut microbiota encode diverse cellulolytic and hemi- tive communities colonizing the microhabitats provided by the cellulolytic enzymes, but the contributions of host and bacterial compartmentalized intestine, including the highly differentiated symbionts to lignocellulose degradation remain ambiguous.
  • Laboratory Exercises in Microbiology: Discovering the Unseen World Through Hands-On Investigation

    Laboratory Exercises in Microbiology: Discovering the Unseen World Through Hands-On Investigation

    City University of New York (CUNY) CUNY Academic Works Open Educational Resources Queensborough Community College 2016 Laboratory Exercises in Microbiology: Discovering the Unseen World Through Hands-On Investigation Joan Petersen CUNY Queensborough Community College Susan McLaughlin CUNY Queensborough Community College How does access to this work benefit ou?y Let us know! More information about this work at: https://academicworks.cuny.edu/qb_oers/16 Discover additional works at: https://academicworks.cuny.edu This work is made publicly available by the City University of New York (CUNY). Contact: [email protected] Laboratory Exercises in Microbiology: Discovering the Unseen World through Hands-On Investigation By Dr. Susan McLaughlin & Dr. Joan Petersen Queensborough Community College Laboratory Exercises in Microbiology: Discovering the Unseen World through Hands-On Investigation Table of Contents Preface………………………………………………………………………………………i Acknowledgments…………………………………………………………………………..ii Microbiology Lab Safety Instructions…………………………………………………...... iii Lab 1. Introduction to Microscopy and Diversity of Cell Types……………………......... 1 Lab 2. Introduction to Aseptic Techniques and Growth Media………………………...... 19 Lab 3. Preparation of Bacterial Smears and Introduction to Staining…………………...... 37 Lab 4. Acid fast and Endospore Staining……………………………………………......... 49 Lab 5. Metabolic Activities of Bacteria…………………………………………….…....... 59 Lab 6. Dichotomous Keys……………………………………………………………......... 77 Lab 7. The Effect of Physical Factors on Microbial Growth……………………………... 85 Lab 8. Chemical Control of Microbial Growth—Disinfectants and Antibiotics…………. 99 Lab 9. The Microbiology of Milk and Food………………………………………………. 111 Lab 10. The Eukaryotes………………………………………………………………........ 123 Lab 11. Clinical Microbiology I; Anaerobic pathogens; Vectors of Infectious Disease….. 141 Lab 12. Clinical Microbiology II—Immunology and the Biolog System………………… 153 Lab 13. Putting it all Together: Case Studies in Microbiology…………………………… 163 Appendix I.
  • Product Sheet Info

    Product Sheet Info

    Product Information Sheet for HM-289 Facklamia sp., Strain HGF4 Incubation: Temperature: 37°C Catalog No. HM-289 Atmosphere: Aerobic Propagation: 1. Keep vial frozen until ready for use, then thaw. For research use only. Not for human use. 2. Transfer the entire thawed aliquot into a single tube of broth. Contributor: 3. Use several drops of the suspension to inoculate an Thomas M. Schmidt, Professor, Department of Microbiology agar slant and/or plate. and Molecular Genetics, Michigan State University, East 4. Incubate the tube, slant and/or plate at 37°C for 48 to Lansing, Michigan, USA 168 hours. Manufacturer: Citation: BEI Resources Acknowledgment for publications should read “The following reagent was obtained through BEI Resources, NIAID, NIH as Product Description: part of the Human Microbiome Project: Facklamia sp., Strain Bacteria Classification: Aerococcaceae, Facklamia HGF4, HM-289.” Species: Facklamia sp. Strain: HGF4 Biosafety Level: 2 Original Source: Facklamia sp., strain HGF4 is a human Appropriate safety procedures should always be used with gastrointestinal isolate.1 this material. Laboratory safety is discussed in the following Comments: Facklamia sp., strain HGF4 (HMP ID 9411) is a publication: U.S. Department of Health and Human Services, reference genome for The Human Microbiome Project Public Health Service, Centers for Disease Control and (HMP). HMP is an initiative to identify and characterize Prevention, and National Institutes of Health. Biosafety in human microbial flora. The complete genome of Facklamia Microbiological and Biomedical Laboratories. 5th ed. sp., strain HGF4 is currently being sequenced at the J. Washington, DC: U.S. Government Printing Office, 2009; see Craig Venter Institute.
  • Abstract: Introduction: Investigation Into The

    Abstract: Introduction: Investigation Into The

    C-109 ASM 106th General Convention - Orlando, Florida - May 22-24, 2006 INVESTIGATION INTO THE EFFECT OF TEMPERATURE ON THE ViaBILITY OF FaSTIDIOUS AND NON-FaSTIDIOUS BaCTERia IN TWO COMMERCiaL AMIES GEL TRANSPORT SYSTEMS: BD CULTURESWAB MaX V(+) AND REMEL BaCTiSWAB. C. BIGGS, THE CHESTER COUNTY HOSPITAL THE CHESTER COUNTY HOSPITAL West Chester, Pennsylvania ABSTRACT: At The Chester County Hospital we use basic insulated cooler boxes with Downingtown, Exton, Kennett Square, and Lionville, Pennsylvania that ice packs for transport of bacteriology swabs from outreach clinics and include various out-patient labs and drawing stations that collect throat physicians offices. During July 2005 we monitored the internal tempera- cultures and urine samples. In addition to this we collect samples from ture of these coolers using digital temperature recorders. Average daily various physicians’ offices. The microbiology lab utilizes a contract cou- temperatures ranged between 11.4 - 19.8º C with peaks as high as 24.3º rier service that is shared with other hospital departments which manages C. We decided to evaluate viability of fastidious bacteria in two Amies Gel the collection of patient samples throughout the day within the network. swab transports; Becton Dickinson CultureSwab Max V(+) (BD) and Transit times for microbiology specimens can vary from 15 minutes Remel BactiSwab (RE) held at room temperature (RT) 20 – 25º C and to several hours depending on timing and courier routing. Before the refrigerator temperature (RF) 4 – 8º C to understand if upgrading our summer of 2005 we had followed guidelines that appear in microbiology coolers to units with electrical refrigeration might improve recovery of reference manuals which advocate the shipment of microbiology swab organisms from swabs.
  • Comparison of the Versatrek® and the Bact/ALERT® Blood Culture Systems for the Growth of Fastidious Microorganisms

    Comparison of the Versatrek® and the Bact/ALERT® Blood Culture Systems for the Growth of Fastidious Microorganisms

    VersaTREK Fastidious Organism Recovery 1 American Society of Microbiology Orlando, Florida - 2005 Poster C-214 Comparison of the VersaTREK® and the BacT/ALERT® Blood Culture Systems for the Growth of Fastidious Microorganisms Fastidious organism recovery in automated blood culture systems is poorly documented. For this reason, in-vitro simulations were performed at Duke University Medical Center comparing the VersaTREK Automated Microbial Detection System (VT) (TREK Diagnostic Systems, Cleveland, OH) to the BacT/ALERT system (3D) (bioMeriéux, Inc., Durham, NC). Thirty seven strains of organisms grown from frozen stocks were used in the study. Organisms were diluted in sterile saline and were targeted to achieve a concentration of 10- 100 CFU/ml. REDOX 1® and REDOX 2® bottles from TREK Diagnostic Systems and BacT/ALERT standard aerobic and anaerobic bottles from bioMeriéux were inoculated with the organisms. The bottles were supplemented with fresh human blood prior to incubation in their respective instrument. To further assess media performance, identical bottles were inoculated with the same organism set and were not supplemented with blood. Parameters assessed in the analysis included recovery of the organisms and mean time to detection in hours for both systems in bottles with and without blood. Results demonstrated the VT REDOX 1 bottle was the most productive bottle in the study. The REDOX 1 bottle (with blood) yielded faster time to detection compared to bioMeriéux standard aerobic bottles for 100% of all aerobic organisms tested. Overall detection for VT was 100% (37/37) compared to 97.0% (36/37) for the 3D system with blood supplementation. The VT system had an overall recovery rate of 94.6% (35/37) compared to 86.5% (32/37) for the 3D system without blood supplementation.
  • CAMP Tests (Standard and Rapid) and Reverse CAMP Test

    CAMP Tests (Standard and Rapid) and Reverse CAMP Test

    M.V.Sc. (Veterinary Microbiology), Monsoon semester Date 31.12.2020 VMC- 602 (Bacteriology II), Unit III, Practical class CAMP Tests (Standard and Rapid) and Reverse CAMP test Dr. Savita Kumari Assistant Professor-cum-Jr. Scientist Department of Veterinary Microbiology Bihar Veterinary College, BASU, Patna CAMP factor S. agalactiae contains the CAMP factor, only beta-hemolytic Streptococcus secretes Pore -forming toxin first identified in this bacterium CAMP reaction is based on the co -hemolytic activity of the CAMP factor Commonly used to identify S. agalactiae Closely related proteins present also in other Gram - positive pathogens cfb gene encodes CAMP factor CAMP test CAMP reaction- consists in a zone of strong hemolysis that is observed when S. agalactiae is streaked next to Staphylococcus aureus on blood agar S. aureus secretes sphingomyelinase Sheep red blood cells - rich in sphingomyelin, and upon exposure to sphingomyelinase become greatly sensitized to CAMP factor, which then effects hemolysis Hemolysis most pronounced in the zone between the colonies of the two bacterial species Co-hemolytic phenomenon- presumptive identification of Group B Streptococci (S. agalactiae) CAMP test First described by Christie, Atkins, and Munch –Petersen in 1944 The protein was named CAMP factor for the initials of the authors of the article that first described the phenomenon Types: Standard CAMP test Rapid CAMP test (spot test ) Standard camp test are time consuming and/or expensive compared to the CAMP spot test Principle CAMP test detects
  • Lactococcus Lactis Cholangitis and Bacteremia Identified by MALDI-TOF Mass Spectrometry

    Lactococcus Lactis Cholangitis and Bacteremia Identified by MALDI-TOF Mass Spectrometry

    J Infect Chemother xxx (2018) 1e6 Contents lists available at ScienceDirect Journal of Infection and Chemotherapy journal homepage: http://www.elsevier.com/locate/jic Case Report Lactococcus lactis cholangitis and bacteremia identified by MALDI-TOF mass spectrometry: A case report and review of the literature on Lactococcus lactis infection* * Akihiko Shimizu a, , Ryota Hase a, b, Daisuke Suzuki a, Akihiro Toguchi c, Yoshihito Otsuka c, Nobuto Hirata d, Naoto Hosokawa a a Department of Infectious Diseases, Kameda Medical Center, 929 Higashicho, Kamogawa, Chiba, 2968602, Japan b Department of Infectious Diseases, Japanese Red Cross Narita Hospital, 90-1 Iidacho, Narita, Chiba, 2868523, Japan c Department of Laboratory Medicine, Kameda Medical Center, 929 Higashicho, Kamogawa, Chiba, 2968602, Japan d Department of Gastroenterology, Kameda Medical Center, 929 Higashicho, Kamogawa, Chiba, 2968602, Japan article info abstract Article history: Lactococcus lactis is a rare causative organism in humans. Cases of L. lactis infection have only rarely been Received 22 March 2018 reported. However, because it is often difficult to identify by conventional commercially available Received in revised form methods, its incidence may be underestimated. We herein report the case of a 70-year-old man with 30 June 2018 cholangiocarcinoma who developed L. lactis cholangitis and review previously reported cases of L. lactis Accepted 17 July 2018 infection. Our case was confirmed by matrix-assisted desorption/ionization time-of-flight mass spec- Available online xxx trometry (MALDI-TOF MS). This case shows L. lactis is a potential causative pathogen of cholangitis and that MALDI-TOF MS can be useful for the rapid and accurate identification of L.
  • Effects of Lactobacillus Rhamnosus and Enterococcus Faecalis Supplementation As Direct-Fed Microbials on Rumen Microbiota of Boer and Speckled Goat Breeds

    Effects of Lactobacillus Rhamnosus and Enterococcus Faecalis Supplementation As Direct-Fed Microbials on Rumen Microbiota of Boer and Speckled Goat Breeds

    veterinary sciences Article Effects of Lactobacillus rhamnosus and Enterococcus faecalis Supplementation as Direct-Fed Microbials on Rumen Microbiota of Boer and Speckled Goat Breeds Takalani Whitney Maake 1,2, Olayinka Ayobami Aiyegoro 2,3,* and Matthew Adekunle Adeleke 1 1 Discipline of Genetics, School of Life Sciences, College of Agricultural, Engineering and Science, University of Kwazulu-Natal, Westville Campus, Private Bag X 54001, Durban 4000, South Africa; [email protected] (T.W.M.); [email protected] (M.A.A.) 2 Gastrointestinal Microbiology and Biotechnology, Agricultural Research Council-Animal Production, Private Bag X 02, Irene 0062, South Africa 3 Research Unit for Environmental Sciences and Management, North-West University, Potchefstroom Campus, Private Bag X 1290, Potchefstroom 2520, South Africa * Correspondence: [email protected] or [email protected]; Tel.: +27-126-729-368 Abstract: The effects on rumen microbial communities of direct-fed probiotics, Lactobacillus rhamnosus and Enterococcus faecalis, singly and in combination as feed supplements to both the Boer and Speckled goats were studied using the Illumina Miseq platform targeting the V3-V4 region of the 16S rRNA microbial genes from sampled rumen fluid. Thirty-six goats of both the Boer and Speckled were divided into five experimental groups: (T1) = diet + Lactobacillus rhamnosus; (T2) = diet + Enterococcus faecalis; (T3) = diet + Lactobacillus rhamnosus + Enterococcus faecalis; (T4, positive control) = diet + antibiotic and (T5, negative control) = diet without antibiotics and without probiotics. Our results revealed that Bacteroidetes, Firmicutes, TM7, Proteobacteria, and Euryarchaeota dominate Citation: Maake, T.W.; Aiyegoro, O.A.; Adeleke, M.A. Effects of the bacterial communities. In our observations, Lactobacillus rhamnosus and Enterococcus faecalis Lactobacillus rhamnosus and supplements reduced the archaeal population of Methanomassiliicocca in the T1, T2 and T3 groups, Enterococcus faecalis Supplementation and caused an increase in the T4 group.