Ocular Infections Caused by Corynebacterium Species

The most well-known Corynebacterium species, Corynebacterium diphtheliae, causes diphthe‐ ria. However, in 1970, the clinical value of identification of Corynebacterium diphtheriae be‐ came less medically significant owing to the development of diphtheria toxoids and a decrease in the prevalence of diphtheritic infection in developed countries. Other Corynebac‐ terium species have been considered contaminants when found in clinical samples because they are organisms normally found in the skin, mucous membranes, and other human tis‐ sues. Given that Corynebacterium species are one of the most commonly isolated bacteria from the ocular surfaces [1, 2], they are also considered non-pathogenic in the ophthalmic field.


Introduction 1.The history of Corynebacterium species
The most well-known Corynebacterium species, Corynebacterium diphtheliae, causes diphtheria.However, in 1970, the clinical value of identification of Corynebacterium diphtheriae became less medically significant owing to the development of diphtheria toxoids and a decrease in the prevalence of diphtheritic infection in developed countries.Other Corynebacterium species have been considered contaminants when found in clinical samples because they are organisms normally found in the skin, mucous membranes, and other human tissues.Given that Corynebacterium species are one of the most commonly isolated bacteria from the ocular surfaces [1,2], they are also considered non-pathogenic in the ophthalmic field.
Currently, in a clinical setting, many bacteriological laboratory technicians in hospitals report Corynebacterium species as "Gram-positive rods".Sometimes, the presence of Corynebacterium species is not reported because it is considered to be contaminants.As a result, it is not possible for ophthalmologists to determine whether Corynebacterium species are present in clinical samples by using laboratory tests.This makes the Corynebacterium species to be nonpathogenic for ophthalmologists leading to therapeutic failure.

Bacteriological characteristics of Corynebacterium species
Morphology: The size of Corynebacterium species varies from 0.3-0.8× 1-8 µm.They exsist in a variety of shapes, even in pure cultures.In the clinical samples, they mostly appear as rod-shaped bacteria in palisade-, ring-, or 'I, N, T, V, W, or Y' letter-shaped arrangments.
Lipophilicity: Few Corynebacterium species generally have high lipophilicity.In vitro, they can be easily become unculturable if the final concentration of Tween 80 (polysorbate 80) in the medium is slightly different from the optimal concentration.Presumably, this is the reason why particular Corynebacterium species prefer the ocular surfaces as these are area where fatty acids are always present because they are secreted from the meibomian gland.This requirement may also explain why Corynebacterium-induced endophthalmitis is very rare.

Corynebacterium species as a pathogen: Case presentations
Case 1: In 2003, the author encountered the case of an elderly patient who had clear infectious conjunctivitis in his right eye.He had experienced mild conjunctival hyperaemia and mucopurulent discharge after cataract surgery performed 2 years before consultation (Fig. 1).He had continued to use a quinolone ophthalmic solution postoperatively, but had not undergone any ophthalmic examination.He had eye discomfort for more than 3 months.Gram staining smear of the discharge showed that many polymorphonuclear leukocytes phagocytizing Gram-positive rods (Fig. 2).Culture of the discharge sample detected quinolone resistant Corynebacterium species, and the strain was susceptible to cephem antibiotics.Switching the quinolone ophthalmic solution to a cephem antibiotic resolved of the patient's symptoms.The author determined this to be a clear case of conjunctivitis due to Corynebacterium species.Thereafter, the author encountered a large number of cases of Corynebacterium conjunctivitis in geriatric patient as well as several cases of Corynebacterium keratitis in patients who underwent keratoplasty.Thus, in 2012, Corynebacterium species still appear to be pathogens of the ocular surface.Case 2: Figures 3 A & B show an ocular surface of a diabetic young man.He had intractable filamentous keratitis after 2 vitrectomies.When he was referred to the author's clinic, a moxifloxacin ophthalmic solution has been prescribed for more than 6 months (from the perioperative stage of the first vitrectomy).After the diagnosis of infectious blepharoconjunctivitis with mucopurulent yellowish discharge, it was determined that the blepharoconjunctivitis may have caused swelling of the eyelid, and the swollen eyelid partially induced intractable filamentous keratitis.Analysis of a smear of the discharge showed a large number of polymorphonuclear leukocytes and Gram-positive rod-shaped bacteria in palisade-and 'I, V, or W' letter-shaped arrangements (Fig. 4).Corynebacterium species were identified in the culture of the discharge by using a simple, commercially available identification kit (BBL Crystal, BD, Japan, Tokyo).The author also isolated Corynebacterium species on a sheep blood agar plate from the discharge and identified the causative agent as Corynebacterium macginleyi on the basis of its biochemical characters tested by API-Coryne (bioMérieux SA, Lyon, France).The minimum inhibitory concentration (MIC) of moxifloxacin and ceftriaxone for the strain (tested by E-test ® , bioMérieux SA, Lyon, France) was >256µg/mL and 2 µg/mL, respectively.Switching moxifloxacin to topical cephmenoxim led to rapid improvement of blepharoconjunctivitis and filamentous keratitis (Fig. 5).
It is currently no exaggeration to say that Corynebacterium species are among the major pathogens responsible for chronic conjunctivitis, especially in geriatric patients.These pathogens can also cause infectious keratitis in patients who are immune-compromised [3][4][5].All such conditions may be triggered, when the bacterial flora of the ocular surface are disturbed, by opportunistic infections.Endophthalmitis caused by Corynebacterium species is very rare.Although C. macginleyi is the common Corynebacterium species to be isolated from the ocular surface [6,7], it remains unclear whether C. macginleyi is the major species responsible for ocular infections because cases caused by other species have been documented as well [5].

Diagnostic techniques
According to Koch's postulates, when establishing the specificity of a pathogenic microorganism, the first criterion is the organism must be present in all cases of the disease.Although quantitative analysis of a specific bacterium in samples by using real-time polymerase chain reaction may be useful, this technique is not readily available to practitioners.It is difficult to validate the other criteria of Koch's postulate, always in clinical setting.Thus, most clinical ophthalmologists depend only on first criterion when identifying a pathogen.
The first step when diagnosing and treating Corynebacterium infections should be to subject the clinical samples, such as mucopurulent discharge or corneal scrapings, to Gram staining, examine them microscopically, and observe whether Gram-positive rods suggestive of Corynebacterium species appear ingested by polymorphonuclear leukocytes (Fig. 2, 4).Finally, the culture results must be accounted.
Although the culture results from discharge and corneal scrapings have clinical significance, we should also recognize the risk of overestimation.As a proof of this, the author has received culture results identifying Staphylococcus epidermidis as a pathogen even though plenty of Gram-positive rods are normally found on microscopy in some patients.In some cases, AB Figure 3. Anterior segments of case 2. Moderate blepharoconjunctivitis, yellowish mucopurulent discharge, and corneal erosion with filamentous keratitis were found.
Infection Control a 'culture negative' result is reported.Figure 6 shows the anterior segments of a bedridden elderly female patient (A) and a panorama Gram stain image of her eye discharge (B).She had a conjunctival hyperaemia with a large amount of yellowish white mucopurulent discharge that lasted for 1 week.The smear prepared from discharge was stained by Gram staining, which showed a large amount of Gram-postitive rods suggestive of Corynebacterium species.Although she clearly had infectious conjunctivitis and no medication had been administered, the culture result from her discharge was reported as negative.Hence, the smear and microscopic examination of clinical samples contribute significantly to the diagnosis of ocular infections caused by Corynebacterium species.

Observation and result
The author found that Corynebacterium species isolated from the ocular surfaces of elderly patients in Japan are very sensitive to cephem antibiotics (Table 1, unpublished data).Although they are also sensitive to aminoglycosides, most of the strains are highly resistant to quinolone [7].

Conclusion
When faced with the case of an elderly patient with chronic conjunctivitis, the first step should be to collect the discharge and to prepare a Gram stained smear and observation under microscope.Assessment should also determine whether the lacrimal duct is obstructed or not.Documenting a patient's history of antimicrobial use will also contribute to the diagnosis.If the patient has a history of using an antimicrobial ophthalmic solution, and also has Gram-positive rods in palisade, ring or 'N, T, V, W, or Y' letter-shaped arrangement present in their discharge and if these Gram positive rods appear to be ingested by polymorphonuclear leukocytes, then a cephem-based ophthalmic solution should be prescribed first.It is possible that an organism other than a Corynebacterium species is the causative pathogen if the cephem antibiotics do not resolve the infection.For Corynebacterium-induced keratitis, a systemic carbapenem and glycopeptide may be useful in additions to frequent applications of cephem, aminoglycoside, and glycopeptide eye drops.

Figure 4 .
Figure 4. Gram stain of the discharge sample from case 2, original magnification ×1000 A large number of Gram-positive rods are phagocytised by polymorphonuclear neutrophil leukocytes.