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Abstract
Introduction: Kocuria kristinae is a commensal organism, sometimes considered as a lab contaminant, but its repeated isolation from clinical samples in immunocompromised patients should raise red flags. Materials and Methods: We confirmed the infection with re-isolation of the organism from the same site before starting treatment. For the identification of Kocuria kristinae we used IDGP cards on VITEK 2 compact system. Antibiotic susceptibility test was done manually following CLSI guidelines 2018 for Coagulase-negative staphylococci. Results: A total of 510 major head neck oncosurgeries were performed during the period of two years. Out of which 120 patients had skin and soft tissue infections. Out of these infected patients, 90 were culture positive and of these Kocuria kristinae were isolated in 12 patients. Resistance to penicillin and oxacillin is seen in all isolates. Conclusion: Kocuria kristinae should not be ignored as a commensal flora or lab contaminant in immunocompromised hosts. Its Increase in resistance pattern is a matter of concern. It is an ignored opportunistic pathogen whose detailed sensitivity test should be developed to treat patients timely and effectively.
Keywords: Head and neck malignancy, Kocuria kristinae, opportunistic pathogen
How to cite this article: Introduction 
Kocuria kristinae is a gram-positive coccus and a normal commensal of skin and oral cavity.[1] In the manual era of identification, it was identified as the only micrococci or coagulase-negative Staphylococcus (CONS).[2] With advances in the automated system, such as VITEK 2 COMPACT, the identification of Kocuria species has become accurate.[3]Kocuria kristinae is reported to cause infections in humans. These are known commensal organisms which can be pathogenic in an immunocompromised host.[4],[5],[6] There are various studies that reported different types of infection in cancer patients under treatment by Kocuria kristinae.[7] Here we are presenting a case series of 12 head and neck cancer patients who developed infection with Kocuria kristinae.
Materials and Methods From May 2019 to May 2021, a total of 510 major head and neck oncosurgeries were performed. Of these, 120 patients had the infection. Pus was collected as routine protocol from the skin or soft tissue of these infected patients.
All patients presented with pus discharge from the skin and underlying soft tissue. There was no differentiating feature between Kocuria kristinae infection and ones caused by other organisms clinically. Aspirate or two sterile cotton swabs or tissue from infected wounds were collected. Normal saline was used to wash the wound prior to the collection of swabs/aspirate. If the wound was dry, then the swab tip was moistened with sterile saline. Care was taken that the swab only came into contact with the wound and not the surrounding skin. Samples were rejected if there was a delay of more than one hour.[8] Ethics Committee approval was taken on 5/7/21.
A smear was prepared from one swab or a drop from aspirate or by smearing the tissue. It was stained by Gram stain and findings were recorded. Another drop of aspirate or second swab or bit of minced tissue was plated on 5% sheep Blood agar (BA) and MacConkey agar (MA). The plates were incubated aerobically at 37°C for 18 to 24 hours. Organisms were identified as gram-positive or gram-negative using Gram stain characteristics. Catalase and coagulase tests were performed.[9]
Organisms from growth were identified as Kocuria Kristinae by VITEK 2 automated system (BioMerieux Inc., Hazelwood, MO, USA) using GPI (Gram-positive identification) card in VITEK 2 COMPACT automated system. Bacitracin disc and furazolidone disc were put to further confirm the diagnosis. Pathogens in all cases were confirmed by taking repeat sample under all aseptic condition and culturing it. As there is no MIC database in the VITEK system for MIC calculation of Kocuria kristinae AST was done by using the database of coagulase-negative staphylococcus and interpretation done as per CLSI guideline for coagulase-negative staphylococcus, 2018. Pigmentation of colonies was further analyzed after 48 hours of incubation.[3],[10],[11],[12]
Due to the limitation of resources we have used bacitracin and furazolidone disc to differentiate it from staphylococci[12] instead of 16s RNA gene sequence analysis.[11]
Results Out of 510 patients which were included in the series, 120 (23.5%) patients presented with skin and soft tissue infection or wound infection during or after their treatment. Of these 120 patients, 90 (75%) patients were culture positive and Kocuria kristinae was identified in 12 (10%) patients. Out of all 12 patients, 11 (91%) were males as shown in [Table 1].
Table 1: Case series of 12 patients with demographic and clinical detailsThe average age of patients who presents with Kocuria infections was 50.8 years (SD 13.5). All infected patients were either stage III or IV.
The average duration of surgery was four hours. In all the surgeries patients received surgical antimicrobial prophylaxis with beta-lactam and beta-lactam inhibitor combination (BLBLI). Average ICU and hospital stay of all patients were 2 and 7 days respectively.
All the patients presented with an infection 42.2 days (SD 51) after surgery. 9 (75%) out of 12 patients had received concurrent chemotherapy and radiotherapy, 1 (8.33%) neo-adjuvant chemotherapy and 1 (8.33%) received concurrent radiotherapy only whereas 1 (8.33%) patient did not receive any therapy before or after surgery.
All the patients were immunocompromised due to tumor cachexia, and/or radiotherapy and chemotherapy. 8 (66%) out of the 12 patients at the time of presentation of infection had leucocytosis with neutrophilia and raised C-Reactive Protein (CRP) levels. Out of 12 patients who developed infection with Kocuria kristinae, only one patient had Diabetes mellitus. No other additional risk factors other than malignancy were known in the rest of the 11 patients.
Out of 12 patients, 6 (50%) patients required readmission. Out of 6 admitted patients, 2 required surgical debridement.
Patients were started on empirical treatment after sending the pus for culture and sensitivity. De-escalation of the therapy was done after a repeat sample was sent to the lab for culture sensitivity to rule out contaminating strains of Kocuria kristinae.
All the patients responded well after de-escalation of therapy to linezolid or vancomycin, with or without fluoroquinolones.
As shown in [Table 2], Gram staining of all clinical samples showed the presence of pus cells and gram-positive cocci. Culture on blood agar after overnight incubation at 37°C showed non hemolytic 0.5-1 mm diameter colonies, opaque and cream-ish white in color, convex with well-defined edges and matted texture as shown in [Figure 1]. On gram staining, these colonies were gram-positive cocci in pairs of tetrads and clusters shown in [Figure 2]. The colonies were catalase-positive and coagulase-negative. VITEK 2 compact identified these colonies as Kocuria kristinae. Colonies were also sensitive to bacitracin and resistant to furazolidone. [Table 3] shows the details of the isolated organisms in all 120 patients who presented with pus discharge.
Table 2: Gram staining and culture sensitivity reports of all isolates of case seriesAntibiotic susceptibility by Kirby Bauer disc diffusion method showed that all isolates (100%) were resistant to penicillin, oxacillin, erythromycin, clindamycin. All isolates (100%) were sensitive to linezolid, daptomycin, quinupristin, and vancomycin. 50% of isolates were sensitive to ciprofloxacin and levofloxacin.
Discussion Kocuria species inhabit the normal skin and mucous membranes of humans and animals.[1],[11],[12],[13] The organism was discovered by Miroslav Kocur, a Slovakian microbiologist. The organism belongs to the family Micrococcaceae and suborder Micrococcineae order Actinomycetales and class Actinobacteria and are gram-positive, aerobic, catalase-positive, coagulase-negative non hemolytic cocci.[11],[12],[13] It is a facultative anaerobe first described in 1974 by Kloos et al. Currently based on the phylogenetic and chemotaxonomic analysis it is classified in genus Kocuria.[14]
Usually regarded as a laboratory contaminant or specimen contaminant it was ignored when isolated from clinical specimens, undermining its pathogenic potential. More than 18 species of Kocuria have been identified based on 16s RNA phylogenetic studies. These are usually considered non-pathogenic bacteria. Recently there is an increase in both superficial and deep-seated infections due to it.[2],[11],[13]
We have used VITEK 2 compact automated system using GPI (Gram-positive identification) card. bacitracin disc and furazolidone disc were put to further confirm the diagnosis.[12],[13]
To our knowledge, this is the biggest case series of infection of Kocuria spp in patients undergoing treatment for malignancy, so very less comparative data is available.
Male predominance is seen in our case series which is also seen by other authors.[7],[12],[15] But in our series that could be a mere coincidence as male predominance is present in head and neck malignancies.[16]
The average age of patients who presented with Kocuria infections was 50.8 years which is not a risk factor as there are variations in age groups in different case series right from neonate to elderly.[4],[5],[6],[7],[11],[13],[15],[17]
Malignancy can be considered as a risk factor or predisposing factor for Kocuria kristinae infections.[7],[12] High rate of infection could be attributed to the site of surgery, late stage of malignancy, tumor cachexia, and long duration of surgery with concurrent chemoradiotherapy.[18]
Eight (66%) out of 12 patients presented with leucocytosis and raised CRP similar to other studies.[7],[13] Further febrile neutropenia, thrombosis, leukocytosis, and elevated CRP are common manifestations of infection with Kocuria.[11] Thus raised CRP and leukocytosis can be used for better clinicopathological correlation with Kocuria infection.
Antimicrobial susceptibility testing done by using CLSI guidelines for CONS was suggested by other authors as well.[7],[12] 100% resistance to penicillin, oxacillin, erythromycin, clindamycin was seen which supports the data of Ahmed et al.[7] The resistance mechanism for penicillin and oxacillin resistance is not clear as proper breakpoints AST criteria are not available.[10],[12]
All isolates (100%) were sensitive to linezolid and vancomycin which again correlates with Ahmed et al.[7] findings. 100% sensitivity to linezolid and high sensitivity to vancomycin is also seen in the review article.[13] In our case series, we did not find resistance to vancomycin. All patients responded well to treatment.
Conclusion The prevalence of human disease caused by kokuria species is underestimated. Kokuria spp has been labeled in the past as a simple contaminant but currently is gaining importance as an emerging pathogen in hosts with cancer, immunocompromised status, and metabolic disorders. Kocuria spp is difficult to identify by conventional method but can be recognized by a modern automated system, which further can be confirmed on 16s RNA phylogenetic studies. Its isolation from clinical specimens should not be ignored.
For testing antibiotic susceptibility there is no proper guideline so there may be chances of misdiagnosis which in turn can lead to recurrent infections and morbid clinical outcomes. The mechanism of drug resistance is least understood. So there is a need for specific criteria for interpreting sensitivity assays with kokuria isolates and deeper investigation of the resistance mechanism. Kokuria species should be studied in detail with genome-based investigations to shed light on the environmental spread, hospital epidemiology, and clinical impact of the pathogen.
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Conflicts of Interest
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References 
Correspondence Address:
Vandana Mohta
21 Kachipura, New Ramdaspeth, Nagpur, Maharashtra - 440010
India
Source of Support: None, Conflict of Interest: None
CheckDOI: 10.4103/ijpm.ijpm_776_21
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