Inducible Clindamycin Resistance (D test)

Clindamycin has been utilized in the treatment of infections caused by Staphylococcus aureus. However, clinical failure to clindamycin therapy is of concern that has been reported due to several mechanisms including resistance to macrolide, lincosamide and streptogramin antibiotics [1]. Routine antibiotic susceptibility test for clindamycin alone may fail to detect inducible clindamycin resistance. In this matter, inducible clindamycin resistance test or D test has resolved the problem so encountered. D test is in fact a simple test that utilizes the two disks clindamycin (2 μg) itself kept in the proximity of erythromycin (15μg) (separated by a distance of 15 mm between the edges). Plates are incubated at 37° C for 24 hr. Inducible resistance to Clindamycin is defined as blunting of the clear circular area of no growth around the Clindamycin disc on the side adjacent to the Erythromycin disc and is designated  as D-test positive. On the contrary to it, absence of a blunted zone of inhibition is designated  as D-test negative. Three different phenotypes are interpreted as MS phenotype, Inducible MLSB phenotype and constitutive MLSB phenotype respectively[2,3] as shown in the table 1.

Methicillin resistant Staphylococcus aureus (MRSA) contributes as a major cause of nosocomial and community acquired infections [4]. The frequency of infection by Methicillin Resistant Staphylococcus aureus (MRSA) has been increased  and has thus altered  the antimicrobial resistance patterns due to appreciable use of macrolide lincosamide-streptogramin B (MLSB) antibiotics to treat such infections [1]. Nevertheless, this has increased the strains resistant to MLSB antibiotics. The MLS family of antibiotics accounts for  three different mechanisms of resistance such as target site modification, enzymatic antibiotic inactivation and macrolide efflux pumps.

Clindamycin is a lincosamide antibiotic which has now become one of the limited choices among antimicrobials effective against MRSA. In the presence of erythromycin resistance, the use of clindamycin in the therapy may led to treatment failures. It is because of the possibility of induction of cross-resistance among members of the macrolide, lincosamide, streptogramin B (MLSB) group [5]. The exaggerated use of clindamycin in both MSSA and MRSA has increased the expression of inducible resistance to clindamycin reducing the effectiveness of this drug. Thus, demonstration of inducible MLSB phenotype in isolates that are susceptible to Clindamycin and resistant to Erythromycin is done by using Double Disk diffusion test or D-test [6,7].

As mentioned earlier, MLSB resistance can be caused by several mechanisms, but the predominant form is target modification mediated by ermA, ermB and ermC (erythromycin ribosome methylase) genes [8,9]. The erm genes encode enzymes that confer inducible or constitutive resistance to MLSB agents via methylation of the 23S rRNA. Thus, it reduces binding by MLSB agents to the ribosome [10,11].

                      Table 1. Different Phenotypes in D-test

Phenotype	Characteristics
1. MS phenotype	Staphylococcal isolates showing circular zone of inhibition around Clindamycin (Zone size > 21mm) and resistance to Erythromycin (Zone size <13 mm)
2. Inducible MLSB phenotype	Staphylococcal isolates showing resistance to Erythromycin (zone size <13 mm) and sensitive to Clindamycin (Zone size>21mm) giving D-shaped zone of inhibition around Clindamycin disk
3. Constitutive MLSB phenotype	Staphylococcal isolates showed resistance to both Erythromycin (Zone size <13 mm) and Clindamycin (Zone size < 14mm) with circular shape of zone of inhibition if any around Clindamycin.

                         

References:
1. Jadhav SV, Gandham N R, Sharma M, Kaur M, Misra R.N. , Matnani G.B. , Ujagare M.T., B. Saikat, Kumar A (2011). Prevalence of inducible Clindamycin resistance among community-and hospital-associated Staphylococcus aureus isolates in a tertiary care hospital in India. Biomedical Research,Vol 22, Issue 4.

2. Deotale V, Mediratta DK, Raut U,et al.Inducible clin-damycin resistance in Staphylococcus aureus isolated from clinical samples.Indain J Med Microbiology 2010; 28 (2): 124-126.

3. Kloos WE, Banerman TL. Staphlococcus and Micro-coccus, Chapter22. In: Manual of clinical microbiol-ogy. 7th ed. Murray PR, Baron EJ, Pfaller MA, Tenoer FC, Yolken RH, editors. Washington DC. ASM Press; 1999: 264-282.

4. Frank AL, Marcinak JF, Mangat PD, et al. Community-acquired methicillin and Clindamycin-susceptible me-thicillin-resistant Staphylococcus aureus in children. Pediatric. Infect. Dis. J 1999; 18: 993-1000.

5. Hussain FM, Boyle-Varva S, Bethel CD, et al. Currents trend in community-acquired methicillin-resistant Sta-phylococcus aureus at a tertiary care pediatric facility. Pediatr. Infect. Dis J 2000; 19: 1163-1166.

6. Gadepalli R, Dhawan B, Mohanthy S, et al. Inducible clindamycin resistance in clinical isolates of Staphylo-coccus aureus. Indian J Med Res 2006; 123: 571-573.


7. Steward CD, Raney PM, Morell AK, et al. Testing for induction of clindamycin resistance in erythromycin re-sistant isolates of Staphylococcus aureus. J Clin Micro-biol 2005; 43: 1716-17121.

8. Leclercq R. Mechanisms of resistance to macrolides and lincosamides:Nature of the resistance elements and their clinical
implications. Clin Infect Dis 2002; 34: 482-492.

9. Roberts MC, Sutcliffe J, Courvalin P, Jensen LB, Rood J, Seppala H. Nomenclature for macrolide-lincosamide-streptogramin B resistance determinants. Antimicrob. Agents

Chemother 1999; 43:2823-2830.

10. Eady EA, Roos JI, Tipper JL, Walters CE, Cove JH, NobleWC. Distribution of genes encoding erythromycin ribosomal methylases and an erythromycin effl ux pump in epidemiologically distinct groups of staphylococci. Antimicrob Agents Chemother 1993;31:211-217.

11. Khan SA, Novick RP. Terminal nucleotide sequences of Tn 551
a transposon specifying erythromycin resistance in Staphylococcus

aureus: homology with Tn3. Plasmid 1980; 4:148-154.