從當前臨床上常見革蘭氏陰性桿菌及陽性球菌對Colistin及Nalidixic acid的藥敏型式探討Columbia CNA agar的效能

Assessing the Efficacy of Columbia CNA Agar Based on the Current Colistin and Nalidixic Acid Susceptibilities of Clinical Gram-Negative Bacilli and Gram-Positive Cocci

林雅筠(Ya-Yun Lin) ； 邱彥昕(Yen-Hsin Chiu) ； 楊榮勝(Rong-Sheng Yang) ； 蔡岳廷(Yueh-ting Tsai) ； 蔡文城(Wen-cherng Tsai)

《檢驗及品保雜誌》 2卷1期 (2013/01) Pp. 1-9

https://doi.org/10.6573/JTQA.201301_2(1).01

摘要

Columbia CNA agar w/5% sheep blood（CNA）為臨床微生物檢驗室針對革蘭氏陽性球菌分離的常用選擇性培養基之一，為了將CNA能更有效地應用於台灣臨床微生物檢驗室，本研究探討臨床上常見革蘭氏陰性桿菌及陽性球菌對colistin及nalidixicacid的藥敏型式，期能找出目前此培養基中之colistin和nalidixicacid兩種抗微生物劑的最適當含量，使CNA在不影響常見革蘭氏陽性球菌的生長下，可抑制高抗藥性革蘭氏陰性桿菌，而增加CNA之應用效能。本研究利用美國臨床及實驗室標準機構（CLSI）出版的方法操作檢測最低抑菌濃度（minimal inhibitory concentration; MIC）的藥敏試驗。檢測時，分別使用含有0.03~64 μg /mL濃度範圍之nalidixicacid以及colistin的Mueller-Hinton agar w/5% sheep blood，以偵測此兩種抗微生物藥劑對各類臨床常見分離菌的MIC。試驗的臨床菌種包括15個菌種（其中5種革蘭氏陽性球菌，10種革蘭氏陰性桿菌），共316個菌株。另外，選擇能在CNA生長的10種革蘭氏陰性桿菌共60株，測定colistin及nalidixicacid對這些菌株的MIC，以了解抗藥程度。操作藥敏試驗時，同時接種Escherichiacoli（ATCC 25922）及Staphylococcus aureus（ATCC 25923）兩個標準菌株作為品管對照，以確認試驗過程的正確性。研究結果指出nalidixic acid對臨床常見革蘭氏陰性桿菌的MIC數值偏高；且當培養基中於的nalidixic acid濃度高於16 μg/mL時會抑制部分革蘭氏陽性球菌的生長。至於colistin濃度，則在其達到10 μg/mL或以上高濃度時，雖然不會抑制臨床上常見的革蘭氏陽性球菌，但仍不能有效地抑制一些革蘭氏陰性桿菌。上述的發現指出，雖然目前含有10 μg/ mL nalidixic acid及10 μg/mL colistin的CNA適合常見革蘭氏陽性球菌的分離，但無法抑制對nalidixic acid及colistin呈高抗藥性的革蘭氏陰性桿菌。為了尋找取代CNA的革蘭氏陽性球菌選擇性培養基，本研究進一步將生長於CNA之10種革蘭氏陰性桿菌，共60個菌株，接種於另一種革蘭氏陽性球菌選擇性培養基：phenylethyl alcohol agar w/5% sheep blood（PEA），結果指出PEA可有效地抑制88.3%生長於CNA之革蘭氏陰性桿菌。總結上述的發現，CNA的配方已經不適合當前台灣臨床微生物檢驗室，但可利用PEA取代CNA作為分離常見的革蘭氏陽性球菌，但仍然要依據blood agar plate（BAP）的生長菌落判讀鏈球菌的溶血型式。

並列摘要

Columbia CNA agar with 5% sheep blood is a Gram-positive cocci selective medium. To make CNA inhibit Gram-negative clinical strains with high drug resistance without affecting the growth of Gram-positive cocci, this study analyzed the susceptibility of clinical Gram-negative bacilli and Gram-positive cocci to colistin and nalidixic acid, with the goal being to find the most appropriate antimicrobial content ratio. In this study, the method published by the Clinical and Laboratory Standards Institute(CLSI) was used to detect the minimum inhibitory concentration (MIC). To determine the MIC of two antimicrobials on various clinical isolates, 0.03-64 μg/mL nalidixic acid and 0.03-64 μg/mL colistin containing Mueller-Hinton (M-H) agar with blood were applied separately. The study tested 316 clinical strains from M-H agar and 60 Gram-negative strains grown on CNA to understand the patterns of drug resistance. The results indicate that nalidixic acid has a high MIC value against clinical Gram-negative bacteria. When the medium contains more than 16 μg/mL of nalidixic acid, it appears to inhibit the growth of Gram-positive cocci. Colistin effectively inhibits most Gram-negative bacteria under 10 μg/mL but does not prevent the growth of Gram-positive bacteria in high antibiotic concentrations. However, for the Gram-negative bacteria grown in CNA, colistin's inhibitory effect is poor even at high concentrations. Therefore, the inhibitory effect test of phenylethyl alcohol (PEA) agar was conducted. The results show that PEA inhibits the growth of 88.3% Gram-negative bacteria grown on CNA. In summary, the CNA formula does not meet the current usage needs of the clinical microbiology laboratories in Taiwan. Alternatively, PEA could replace CNA to isolate Gram-positive cocci, but the hemolytic-type interpretation of Streptococcus should still depend on the colonies' growth on blood agar plates.

並列關鍵字

Columbia CNA agar ； Nalidixic acid ； Colistin ； MIC ； PEA