460
Indian Journal of Medical Microbiology
aeruginosa PAO1 strain in this study. All isolates were tested for neuraminidase production by sensitive fluorometric assay using 2’‑(4‑methylumbelliferyl)α‑D‑N acetylneuraminic acid (Sigma‑Aldrich, USA) as substrate.[7] 47/108 (43.5%) isolates of A. baumannii from clinical specimens were found to produce neuraminidase, only 8 (28.5%) isolates (5 A. lwoffii and 3 A. junii) were producing neuraminidase from 28 (24 A. lwoffii and 4 A. junii) isolates of skin of healthy individuals. Similarly study conducted by Sugita et al. in 2000 documented that 2‑3% of Acinetobacter spp were found to have neuraminidase activity.[6] Further, these isolates were also tested for biofilm production by tissue culture plate method.[8] Among neuraminidase‑positive isolates, 19/47 (40.4%) clinical isolates of A. baumannii and 3/8 (37.5%) skin isolates were shown to form biofilm. Whereas, 12/61 (19.6%) and 6/20 (30%) of neuraminidase negative clinical isolates of A. baumannii and skin isolates from healthy individuals, respectively, were found to exhibit biofilm. Neuraminidase producing these bacteria can convert sialyated to asialyated glycolipids on the mucosal surfaces, facilitating the attachment of such bacteria on to the host cells followed by colonisation.[4] 40.4% of neuraminidase‑positive A. baumannii clinical isolates and 37.5% of skin neuraminidase‑positive isolates were biofilm producers. We concluded that neuraminidase enzyme can also be one of the contributing virulent factor of Acinetobacter spp for the attachment towards various surfaces. Further study involving with molecular mechanism of neuraminidase exhibited by Acinetobacter spp may throw more light on this virulence trait to know the actual pathogenesis of the outrageous pathogen. This is the first and preliminary study conducted on this factor in Indian context.
4.
5.
6. 7.
8.
vol. 32, No. 4
susceptibility of Acinetobacter species on the skin of healthy humans. Natl Med J India 2001;14:204‑8. Soong G, Muir A, Gomez MI, Waks J, Reddy B, Planet P, et al. Bacterial neuraminidase facilitates mucosal infection by participating in biofilm production. J Clin Invest 2006;116:2297‑305. Cacalano G, Kays M, Saiman L, Prince A. Production of Pseudomonas aeruginosa neuraminidase is increased under hyper osmolar conditions and is regulated by genes involved in alginate expression. J Clin Invest 1992;89:1866‑74. Sugita H, Shinagawa Y, Okano R. Neuraminidase producing ability of intestinal bacteria isolated from coastal fish. Lett Appl Microbiol 2000;31:10‑3. Ghazei C, Ahmadi M, Jazani NH. Detection of neuraminidase activity in Pseudomonas aeruginosa PAO1. Iran J Basic Med Sci 2010;13:69-5. Mathur T, Singhal S, Khan S, Upadhyay DJ, Fatma T, Rattan A. Detection of biofilm formation among the clinical isolates of Staphylococci: An evaluation of three different screening methods. Indian J Med Microbiol 2006;24:25‑9.
Gopinath Prakasam, Anusha Rohit, E Padmasini, *S Srivani Ramesh Department of Microbiology (GP, P, SSR), Dr. Arcot Lakshmanaswami Mudaliar Post Graduate Institute of Basic Medical Sciences, University of Madras, Taramani, Chennai, Tamil Nadu, India Department of Clinical Microbiology (AR), Madras Medical Mission, Mogappair, Chennai, Tamil Nadu, India. *Corresponding author (email: ) Received: 30‑09‑2013 Accepted: 06-02-2014
References
Access this article online
1. Peleg AY, Seifert H, Paterson DV. Acinetobacter baumannii: Emergence of a successful pathogen. Clin Microbiol Rev 2008;21:538‑82. 2. Patil JR, Jog NR, Chopade BA. Isolation and characterization of Acinetobacter species from upper respiratory tract of healthy humans and demonstration of lectin activity. Indian J Med Microbiol 2001;19:30‑5. 3. Patil JR, Chopade BA. Distribution and in vitro antimicrobial
Quick Response Code:
Website: www.ijmm.org PMID: *** DOI: 10.4103/0255-0857.142235
Small colony variants of Staphylococcus aureus: Enemies with hidden weapons
Dear Editor, Small colony variants (SCV) of Staphylococcus aureus originate from wild‑type strains by genetic mutation, resulting in loss of properties like α‑cytotoxin (α‑hemolysin) activity and auxotrophicity for thymidine
and menadione or vitamin K.[1] These morphological variants are typified by a slow growth rate, absence of colony pigment, colony size approximately one‑tenth that of wild‑type strains and atypical biochemical characteristics, which make laboratory diagnosis very difficult.[2]
www.ijmm.org
October - December 2014
461
Correspondences
SCVs are auxotrophic for menadione and thymidine. Intra‑cellular persistence shields them from host immunity and antibiotics.[3] The differentiating features of SCV and wild‑type S. aureus are enumerated in Table 1.[4] SCVs are associated with diseases like cystic fibrosis, osteomyelitis and wound infection.[5,6] SCV of S. aureus can also be recovered in 17-49% cases of cystic fibrosis. There are also reports of device‑related infections, like ventriculo‑peritoneal shunt infections. Diagnosis follows timely suspicion since colonies are slow‑growing, and multiply at a rate one‑ninth that of wild‑type S. aureus. Accurate diagnosis rests upon detection of small, pinpoint, non‑pigmented, non‑haemolytic colonies in 5% Sheep blood agar reverting to normal phenotype in presence of vitamin K in media (Schaedler agar) and ambient 5% CO2. “Fried Egg” colonies (central raised and peripheral flat parts) are characteristic of Thymidine‑auxotrophic SCV. Colonies are also catalase positive and delayed positive for tube coagulase. However, tube coagulase comes positive only after 18 h incubation.Molecular methods like detection of S. aureus‑specific nuc or coa genes (by polymerase chain reaction) can also be attempted. Some authors have shown augmented growth on lawn culture around discs impregnated with 15 µl of vitamin K as diagnostic. Colonies of SCV S. aureus have been shown in Figure 1. Table 1: Differences between wild type S. aureus and SCV Property Wild type SCV Growth Fast (48 h Pigment Present Absent Colony size 2-3 mm 0.2-0.3 mm Coagulase Fast positive Slow positive Mannitol fermentation +ve −ve Haemolysis Present Absent
Treatment is difficult due to inherent resistance to aminoglycosides and cell‑wall active antibiotics. Rifampicin, being active intra‑cellularly, or its combination with cotrimoxazole is effective. Daptomycin injection for about 2 months is documented to be useful in deep‑seated infections.[7] SCVs of S. aureus represent underdiagnosed agents of persistent subclinical infections that need to be promptly diagnosed and treated. Further research is awaited regarding their pathogenesis and treatment guidelines. The authors thank all workers whose research has been cited and mentioned in this study. References 1. Melter O, Radojevic B. Small colony variants of Staphylococcus aureus‑‑review. Folia Microbiol (Praha) 2010;55:548‑58. 2. Besier S, Smaczny C, von Mallinckrodt C, Krahl A, Ackermann H, Brade V, et al. Prevalence and clinical significance of Staphylococcus aureus small‑colony variants in cystic fibrosis lung disease. J Clin Microbiol 2007;45:168‑72. 3. Sifri CD, Baresch‑Bernal A, Calderwood SB, von Eiff C. Virulence of Staphylococcus aureus small colony variants in the Caenorhabditis elegans infection model. Infect Immun 2006;74:1091‑6. 4. Atalla H, Gyles C, Mallard B. Staphylococcus aureus small colony variants (SCVs) and their role in disease. Anim Health Res Rev 2011;12:33‑45. 5. Yagci S, Hascelik G, Dogru D, Ozcelik U, Sener B. Prevalence and genetic diversity of Staphylococcus aureus small‑colony variants in cystic fibrosis patients. Clin Microbiol Infect 2013;19:77‑84. 6. Proctor RA, van Langevelde P, Kristjansson M, Maslow JN, Arbeit RD. Persistent and relapsing infections associated with small‑colony variants of staphylococcus aureus. Clin Infect Dis 1995;20:95‑102. 7. Kipp F, Ziebuhr W, Becker K, Krimmer V, Höbeta N, Peters G, et al. Detection of Staphylococcus aureus by 16S rRNA directed in situ hybridisation in a patient with a brain abscess caused by small colony variants. J Neurol Neurosurg Psychiatry 2003;74:1000‑2.
SCV: Small colony variants
*S Bhattacharyya, D Kumar Department of Microbiology (SB), All India Institute of Medical Sciences, Phulwarisharif, Patna ‑ 801 505, Bihar, Department of Microbiology, IMS, BHU, Varanasi, Uttar Pradesh, India. *Corresponding author: (email: ) Received: 05‑09‑2013 Accepted: 20‑01‑2014 Access this article online Quick Response Code:
Website: www.ijmm.org PMID: *** DOI: 10.4103/0255-0857.142237
Figure 1: SCV colonis on 5% sheep blood agar www.ijmm.org
Copyright of Indian Journal of Medical Microbiology is the property of Medknow Publications & Media Pvt. Ltd. and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use.
Indian Journal of Medical Microbiology
aeruginosa PAO1 strain in this study. All isolates were tested for neuraminidase production by sensitive fluorometric assay using 2’‑(4‑methylumbelliferyl)α‑D‑N acetylneuraminic acid (Sigma‑Aldrich, USA) as substrate.[7] 47/108 (43.5%) isolates of A. baumannii from clinical specimens were found to produce neuraminidase, only 8 (28.5%) isolates (5 A. lwoffii and 3 A. junii) were producing neuraminidase from 28 (24 A. lwoffii and 4 A. junii) isolates of skin of healthy individuals. Similarly study conducted by Sugita et al. in 2000 documented that 2‑3% of Acinetobacter spp were found to have neuraminidase activity.[6] Further, these isolates were also tested for biofilm production by tissue culture plate method.[8] Among neuraminidase‑positive isolates, 19/47 (40.4%) clinical isolates of A. baumannii and 3/8 (37.5%) skin isolates were shown to form biofilm. Whereas, 12/61 (19.6%) and 6/20 (30%) of neuraminidase negative clinical isolates of A. baumannii and skin isolates from healthy individuals, respectively, were found to exhibit biofilm. Neuraminidase producing these bacteria can convert sialyated to asialyated glycolipids on the mucosal surfaces, facilitating the attachment of such bacteria on to the host cells followed by colonisation.[4] 40.4% of neuraminidase‑positive A. baumannii clinical isolates and 37.5% of skin neuraminidase‑positive isolates were biofilm producers. We concluded that neuraminidase enzyme can also be one of the contributing virulent factor of Acinetobacter spp for the attachment towards various surfaces. Further study involving with molecular mechanism of neuraminidase exhibited by Acinetobacter spp may throw more light on this virulence trait to know the actual pathogenesis of the outrageous pathogen. This is the first and preliminary study conducted on this factor in Indian context.
4.
5.
6. 7.
8.
vol. 32, No. 4
susceptibility of Acinetobacter species on the skin of healthy humans. Natl Med J India 2001;14:204‑8. Soong G, Muir A, Gomez MI, Waks J, Reddy B, Planet P, et al. Bacterial neuraminidase facilitates mucosal infection by participating in biofilm production. J Clin Invest 2006;116:2297‑305. Cacalano G, Kays M, Saiman L, Prince A. Production of Pseudomonas aeruginosa neuraminidase is increased under hyper osmolar conditions and is regulated by genes involved in alginate expression. J Clin Invest 1992;89:1866‑74. Sugita H, Shinagawa Y, Okano R. Neuraminidase producing ability of intestinal bacteria isolated from coastal fish. Lett Appl Microbiol 2000;31:10‑3. Ghazei C, Ahmadi M, Jazani NH. Detection of neuraminidase activity in Pseudomonas aeruginosa PAO1. Iran J Basic Med Sci 2010;13:69-5. Mathur T, Singhal S, Khan S, Upadhyay DJ, Fatma T, Rattan A. Detection of biofilm formation among the clinical isolates of Staphylococci: An evaluation of three different screening methods. Indian J Med Microbiol 2006;24:25‑9.
Gopinath Prakasam, Anusha Rohit, E Padmasini, *S Srivani Ramesh Department of Microbiology (GP, P, SSR), Dr. Arcot Lakshmanaswami Mudaliar Post Graduate Institute of Basic Medical Sciences, University of Madras, Taramani, Chennai, Tamil Nadu, India Department of Clinical Microbiology (AR), Madras Medical Mission, Mogappair, Chennai, Tamil Nadu, India. *Corresponding author (email: ) Received: 30‑09‑2013 Accepted: 06-02-2014
References
Access this article online
1. Peleg AY, Seifert H, Paterson DV. Acinetobacter baumannii: Emergence of a successful pathogen. Clin Microbiol Rev 2008;21:538‑82. 2. Patil JR, Jog NR, Chopade BA. Isolation and characterization of Acinetobacter species from upper respiratory tract of healthy humans and demonstration of lectin activity. Indian J Med Microbiol 2001;19:30‑5. 3. Patil JR, Chopade BA. Distribution and in vitro antimicrobial
Quick Response Code:
Website: www.ijmm.org PMID: *** DOI: 10.4103/0255-0857.142235
Small colony variants of Staphylococcus aureus: Enemies with hidden weapons
Dear Editor, Small colony variants (SCV) of Staphylococcus aureus originate from wild‑type strains by genetic mutation, resulting in loss of properties like α‑cytotoxin (α‑hemolysin) activity and auxotrophicity for thymidine
and menadione or vitamin K.[1] These morphological variants are typified by a slow growth rate, absence of colony pigment, colony size approximately one‑tenth that of wild‑type strains and atypical biochemical characteristics, which make laboratory diagnosis very difficult.[2]
www.ijmm.org
October - December 2014
461
Correspondences
SCVs are auxotrophic for menadione and thymidine. Intra‑cellular persistence shields them from host immunity and antibiotics.[3] The differentiating features of SCV and wild‑type S. aureus are enumerated in Table 1.[4] SCVs are associated with diseases like cystic fibrosis, osteomyelitis and wound infection.[5,6] SCV of S. aureus can also be recovered in 17-49% cases of cystic fibrosis. There are also reports of device‑related infections, like ventriculo‑peritoneal shunt infections. Diagnosis follows timely suspicion since colonies are slow‑growing, and multiply at a rate one‑ninth that of wild‑type S. aureus. Accurate diagnosis rests upon detection of small, pinpoint, non‑pigmented, non‑haemolytic colonies in 5% Sheep blood agar reverting to normal phenotype in presence of vitamin K in media (Schaedler agar) and ambient 5% CO2. “Fried Egg” colonies (central raised and peripheral flat parts) are characteristic of Thymidine‑auxotrophic SCV. Colonies are also catalase positive and delayed positive for tube coagulase. However, tube coagulase comes positive only after 18 h incubation.Molecular methods like detection of S. aureus‑specific nuc or coa genes (by polymerase chain reaction) can also be attempted. Some authors have shown augmented growth on lawn culture around discs impregnated with 15 µl of vitamin K as diagnostic. Colonies of SCV S. aureus have been shown in Figure 1. Table 1: Differences between wild type S. aureus and SCV Property Wild type SCV Growth Fast (48 h Pigment Present Absent Colony size 2-3 mm 0.2-0.3 mm Coagulase Fast positive Slow positive Mannitol fermentation +ve −ve Haemolysis Present Absent
Treatment is difficult due to inherent resistance to aminoglycosides and cell‑wall active antibiotics. Rifampicin, being active intra‑cellularly, or its combination with cotrimoxazole is effective. Daptomycin injection for about 2 months is documented to be useful in deep‑seated infections.[7] SCVs of S. aureus represent underdiagnosed agents of persistent subclinical infections that need to be promptly diagnosed and treated. Further research is awaited regarding their pathogenesis and treatment guidelines. The authors thank all workers whose research has been cited and mentioned in this study. References 1. Melter O, Radojevic B. Small colony variants of Staphylococcus aureus‑‑review. Folia Microbiol (Praha) 2010;55:548‑58. 2. Besier S, Smaczny C, von Mallinckrodt C, Krahl A, Ackermann H, Brade V, et al. Prevalence and clinical significance of Staphylococcus aureus small‑colony variants in cystic fibrosis lung disease. J Clin Microbiol 2007;45:168‑72. 3. Sifri CD, Baresch‑Bernal A, Calderwood SB, von Eiff C. Virulence of Staphylococcus aureus small colony variants in the Caenorhabditis elegans infection model. Infect Immun 2006;74:1091‑6. 4. Atalla H, Gyles C, Mallard B. Staphylococcus aureus small colony variants (SCVs) and their role in disease. Anim Health Res Rev 2011;12:33‑45. 5. Yagci S, Hascelik G, Dogru D, Ozcelik U, Sener B. Prevalence and genetic diversity of Staphylococcus aureus small‑colony variants in cystic fibrosis patients. Clin Microbiol Infect 2013;19:77‑84. 6. Proctor RA, van Langevelde P, Kristjansson M, Maslow JN, Arbeit RD. Persistent and relapsing infections associated with small‑colony variants of staphylococcus aureus. Clin Infect Dis 1995;20:95‑102. 7. Kipp F, Ziebuhr W, Becker K, Krimmer V, Höbeta N, Peters G, et al. Detection of Staphylococcus aureus by 16S rRNA directed in situ hybridisation in a patient with a brain abscess caused by small colony variants. J Neurol Neurosurg Psychiatry 2003;74:1000‑2.
SCV: Small colony variants
*S Bhattacharyya, D Kumar Department of Microbiology (SB), All India Institute of Medical Sciences, Phulwarisharif, Patna ‑ 801 505, Bihar, Department of Microbiology, IMS, BHU, Varanasi, Uttar Pradesh, India. *Corresponding author: (email: ) Received: 05‑09‑2013 Accepted: 20‑01‑2014 Access this article online Quick Response Code:
Website: www.ijmm.org PMID: *** DOI: 10.4103/0255-0857.142237
Figure 1: SCV colonis on 5% sheep blood agar www.ijmm.org
Copyright of Indian Journal of Medical Microbiology is the property of Medknow Publications & Media Pvt. Ltd. and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use.
