Lab Matters Summer 2018 - Page 25

public health preparedness and response APHL Updates Resources and Outreach for Sentinel Clinical Laboratories By Samuel Abrams, MPH, specialist, Public Health Preparedness and Response Biological threat (BT) coordinators are tasked with ensuring that thousands of clinical laboratories across the country are adequately prepared to properly rule-out and refer threats. The benefits of training are obvious for the safety of both the patient and the lab scientist, but outreach can be an arduous task. To address this need, APHL coordinated the update and release of guidance resources to help public health laboratories connect with sentinel clinical laboratories. Representatives from state and local public health laboratories, the American Society for Microbiology (ASM) and the US Centers for Disease Control and Prevention (CDC) participated in the development of the materials. Earlier this year, updates were made to the Biothreat Agent Bench Cards, a bench-side flipchart that provides a quick reference point for some select agents. The inclusion of identification methods for B. cereus biovar anthracis and a Gram- negative bacilli/coccobacilli rule-out flowchart were two valuable additions. Other updates included alignment with ASM sentinel level clinical laboratory protocols. Furthermore, the bench cards provide new information for biosafety, including details on performing biological risk assessments. An accompanying wall-mounted poster (see below) was also updated, outlining the select agents’ characterization details. Both of these resources were sent out via email to all APHL members, BT Coordinators and Biosafety officers in June, 2018. Another major update has been to the sentinel clinical laboratory definition. The changes provide flexibility for state public health laboratories to work with their own non-microbiological sentinel laboratories for outreach and training. The definition addresses moderate complexity testing by including statements on point- of-care testing and culture-independent diagnostic tests, providing guidance for a range of testing sites that previously have not been included on guidance- related issues. The update also includes statements on biosafety and lays out the responsibilities of core laboratories to communicate important biothreat information with satellite laboratories, ensuring that critical information is clearly outlined and transmitted in a timely manner. APHL also supports training needs assessments and preparedness training programs for public health and clinical laboratories. n BIOTHREAT AGENTS ANTHRAX Bacillus anthrac is • Large Gram positive rods (1-1.5 µm x 3-5 µm) • Smears of clinical specimens: • Short chains (2-4 cells) • Capsule present, no spores • Smears from BAP and CHOC culture: • Long chains, no capsule • Spores in older cultures; oval, central to subterminal, no swelling of cell wall • Grows well on BAP and CHOC • No growth on MAC and EMB • Ground-glass colonies, 2-5 mm on BAP and CHOC at 24h • Aerobic growth as early as 4-8h • Flat or slightly convex with irregular edges that may have comma-like projections BRUCELLOSIS GLANDERS Brucella spp. • Tiny, faintly staining, non- clustered, Gram negative coccobacilli (0.4 µm-0.8 µm) • Pinpoint colonies at 24h, and 0.5-1.0 mm after 48h • Non-hemolytic MELIOIDOSIS Burkholderia mallei Burkholderia pseudomallei • Small straight, or slightly curved with rounded ends, Gram negative coccobacilli (1.5 µm-3 µm x 0.5-1.0 µm) • Straight, or slightly curved Gram negative rod (2.0-5.0 µm x 0.4-0.8 µm) • Cells arranged in pairs, parallel bundles, or Chinese letter form • Colonies may demonstrate bipolar morphology in direct specimens and peripheral staining in older cultures, which can mimic endospores • Non-mucoid • Aerobic • Aerobic growth on BAP and CHOC (CO 2 may be required by some strains) • Non-hemolytic • No growth or pinpoint on MAC at 48h • No growth on MAC or EMB • Catalase positive • Catalase, oxidase, urea: positive (Oxidase may be variable) • Oxidase variable • X and V factor (satellite test) negative (not required) • Non-motile • Distinctive musty earthy odor, which is diagnostic (the odor is apparent without sniffi ng) • No growth at 42°C • Oxidase positive • Polymyxin B and colistin no zone • Spot indole negative • Penicillin resistant • Motile • Amoxicillin-clavulanate susceptible • Growth at 42°C • Non-motile (although motility testing not recommended for suspect Brucella spp.) • Non-hemolytic on BAP • Spot indole negative • Aerobic • Non-hemolytic • Growth on MAC (may uptake pink dye) • Polymyxin B and colistin no zone • Penicillin resistant • Tenacious, sticky colonies, adheres to agar surface • Amoxicillin-clavulanate susceptible • Catalase positive TULAREMIA PLAGUE Francisella tularensis Yersinia pestis • Tiny, Gram negative coccobacilli (0.2-0.5 µm x 0.7-1.0 µm) • Plump, Gram negative rods (0.5 x 1-2 µm) seen mostly as single cells or pairs, and may demonstrate short chains in liquid media • Poor counterstaining with safranin (basic fuchsin counter- stain may increase resolution) • May exhibit bipolar, “safety-pin” appearance in Giemsa stain or Wright’s stain • Pleomorphic • Mostly single cells • Aerobic, fastidious • Facultative anaerobe • No growth on MAC/EMB • Scant or no growth on BAP; may grow on primary culture, not well on subculture • Slow growing at 35˚C, better growth at 25-28˚C • Slow growing on CHOC, TM or BCYE: 1-2 mm after 48h • Grey-white, translucent pinpoint colonies at 24h, usually too small to be seen, little to no hemolysis on BAP • Colonies are opaque, grey-white, butyrous, smooth and shiny • At 48h, lactose non-fermenter on MAC or EMB • Oxidase negative • Catalase positive • Catalase negative or weakly positive • Oxidase, urease (at 35˚C) and indole negative • Satellite negative • Beta-lactamase positive • Non-motile FOLLOW ALL LABORATORY AND BIOSAFETY PROCEDURES TO RECOGNIZE AGENTS OF BIOTERRORISM YOU ARE THE FIRST LINE OF DEFENSE — REFER TO CURRENT ASM SENTINEL LAB PROTOCOLS PublicHealthLabs @APHL ® Summer 2018 LAB MATTERS 23