Toll-like receptors in acute post-cataract surgery endophthalmitis.

OBJECTIVE: To evaluate the expression of toll-like receptor 2 (TLR2) and toll-like receptor 4 (TLR4) on CD14 + cells in vitreous and blood of post-cataract surgery acute endophthalmitis. DESIGN: This prospective case-control pilot study enrolled 16 patients of post-cataract surgery endophthalmitis. All the cases were subjected to 23 G pars plana vitrectomy (PPV). Ten patients undergoing 23 G PPV for non-infectious conditions were taken as controls. METHODS: 23 G PPV was performed, and three undiluted vitreous samples were collected in heparinized syringes from the cases and the controls. Simultaneous venous blood sample was taken, and flow cytometry was performed to detect the expression of TLR2 and TLR4 in vitreous and blood samples. The vitreous and blood samples were incubated with fluorescein isothicyanate (FITC) conjugated anti-TLR2 monoclonal antibody Alexafluor (AX) 647 and anti-TLR4 monoclonal antibody phycoerythrin. Data acquisition was done on a pre-calibrated flow cytometer. TLR analysis of the acquired flow cytometry data was then performed. Mean channel fluorescence intensity (MFI) derived from fluorescence histogram was used to study the level of cell surface TLR expression. MFI was calculated as a ratio and recorded as the MFI of the TLR2 or -4 antibody divided by the MFI of the isotype-matched negative control antibody. Core vitrectomy was done as per the comfort of the surgeon, and intravitreal antibiotics vancomycin (1 mg/0.1 ml) and ceftazidime (2.25 mg/0.1 ml) were injected. The cytological examination was done on vitreous and blood sample. STATISTICAL ANALYSIS: The median TLR 2 and TLR4 values between cases and controls were compared by Mann-Whitney U test. Spearman's rank correlation test was used to assess the correlation between TLR expression and disease activity. RESULTS: Vitreous cytology evaluation showed the presence of neutrophils (81.25%, n = 13), monocytes (68.75%, n = 11) and lymphocytes (62.50%, n = 10). The level of expression of TLR2 in vitreous showed a statistically significant correlation with an increase in the time interval of cataract surgery and intervention for endophthalmitis (p < 0.05), but the same was not observed for TLR4. A drift toward higher level of expression of TLR2 and TLR4 in vitreous was observed in patients with poor outcome. CONCLUSION: TLR2 levels increase with the delay in presentation; thus, TLR2 ligands in vitreous could serve as a good target for the treatment of endophthalmitis.

Two cases of primary vitreoretinal lymphoma: a diagnostic challenge : The supporting role of multimodal imaging in the diagnosis of primary vitreoretinal lymphoma.

PURPOSE: To report two cases of primary vitreoretinal lymphoma (PVRL), which presented as intermediate and posterior uveitis. METHODS: Combined clinical assessment, multimodal imaging with spectral-domain optical coherence tomography, fundus autofluorescence, fluorescein angiography, indocyanine green angiography, brain magnetic resonance imaging and vitreous and retinal biopsy. Case 1 was a 48-year-old woman who complained of visual loss in her right eye secondary to a diffuse vitreous opacification and multiple chorioretinal lesions. Case 2, a 74-year-old man, presented with low vision in his right eye due to a wide chorioretinal lesion at the posterior pole, vitreous opacification and posterior uveitis in both eyes. RESULTS: Diffuse large B cell lymphoma was histologically diagnosed in the cerebellum in the first case and in chorioretinal tissue in the second patient. Atypical lymphoid cells were detected and allowed to make a diagnosis of primary central nervous system lymphoma in case 1 and PVRL in case 2. CONCLUSION: PVRL often masquerades ad intermediate or posterior uveitis. The management of the patients needed a team of pathologists, haematologists and ophthalmologists to achieve the correct diagnosis and choose the more appropriate therapy. Some peculiar characteristics on multimodal imaging, even in atypical cases of PVRL, should raise suspicious for PVRL and lead to a diagnostic vitrectomy and/or retinal biopsy.

Electroretinographic and Optical Coherence Tomographic Evaluations of Eyes with Vitreoretinal Lymphoma.

Vitreoretinal lymphomas (VRLs) present with different clinical characteristics. However, only a few case reports have been published that evaluated the retinal function and the retinal morphology. The relationship between retinal morphology and function of eyes with a vitreoretinal lymphoma (VRL) was investigated via optical coherence tomography (OCT) and electroretinography (ERG). The ERG and OCT findings in 11 eyes of 11 patients (69.4 ± 11.5 years old) who were diagnosed with VRL at the Saitama Medical University Hospital between December 2016 to May 2022 were studied. The decimal best-corrected visual acuity ranged from hand movements to 1.2 (median 0.2). Histopathological studies of the vitreous specimens showed class II VRL in one eye, class III VRL in seven eyes, class IV VRL in two eyes, and class V VRL in one eye. The IgH gene rearrangement was positive in three of the six eyes tested. The OCT images showed morphological abnormalities in 10 of the 11 (90.9%) eyes. Severe attenuation was found for the amplitudes of the b-wave of the DA 0.01 ERG in 6 of 11 eyes (54.5%), the DA 3.0 a-wave in 5 of 11 eyes (45.5%), the DA 3.0 b-wave in 36.4%, the LA 3.0 a-wave in 36.4%, the LA 3.0 b-wave in 18.2%, and flicker responses in 36.4% of the eyes. None of the DA 3.0 ERGs had a negative shape (b/a < 1.0). In the five eyes in which the a-wave was severely attenuated, hyperreflective dots were observed subretinally. The ERG analysis in eyes with a VRL indicates a relatively severe dysfunction of the outer retinal layer and was helpful in determining the site of the morphological changes in eyes with VRL.

Laboratory Investigations in Infectious Uveitis.

Laboratory investigations can play a significant role in the diagnosis and decision-making of infectious uveitis. Though direct demonstration of the infective organism remains the gold standard of diagnosis, it is not always possible with ocular tissues. Recent advancements in molecular techniques have made it possible to overcome these limitations and to identify the genomic DNA of pathogens associated with infectious uveitis. Techniques such as next-generation sequencing can analyze all DNA-based lifeforms, regardless of whether they are bacteria, fungi, viruses, or parasites and have been used in the laboratory diagnosis of intraocular inflammation. On the other hand, serological tests, though they dominate the diagnostic landscape of various infectious etiologies in uveitis in routine clinical practice, have varied specificities and sensitivities in different infectious uveitis. In this review, we focus on various methods of laboratory diagnosis of infectious uveitis and discuss the recent advances in molecular diagnosis and their role in various infectious clinical entities.

Molecular Analysis of Liquid Vitreous Biopsy Reveals Occult Lymphoma Following Cytology-Negative Biopsies of the Brain and Vitreous.

PURPOSE: Primary central nervous system lymphoma (PCNSL) is a rare but deadly malignancy that principally affects adults in the fifth and sixth decades of life. Despite diagnostic advances in analyses of cerebral spinal fluid and neuroimaging, definitive diagnosis of PCNSL requires primary brain tissue biopsy. While small neurosurgical biopsy volumes are pursued to minimize removal of normal brain tissue, the spatial margins to precisely biopsy pathologic tissue are narrow and can result in missed diagnoses. Furthermore, prior steroid treatment can significantly reduce tumor burden increasing the likelihood of a non-diagnostic biopsy. METHODS: A retrospective case report from a tertiary referral center using a combination of neuroradiological studies, sterotactic tissue biopsy, and molecular testing for genome mutations. RESULTS: A 72-year-old woman with strong suspicion for PCNSL clinically and radiologically, but cerebral spinal fluid and primary brain tissue biopsy were negative for tumor. However, vitreous liquid biopsy molecular testing for a MYD88 mutation as well as B-cell clonality (IGH/IGK rearrangement) were positive, indicating the presence of secondary vitreoretinal lymphoma from PCNSL. Only after autopsy of her brain was histopathological and immunohistochemical evidence of PCNSL confirmed. CONCLUSION: This case illustrates the unique contribution of liquid biopsy neuropathology-oriented molecular testing in a challenging case with high clinical suspicion of PCNSL in which gold-standard diagnostic testing failed to yield a diagnosis.

Diagnostic sensitivity of vitrectomy and vitreous biopsy for vitritis.

PURPOSE: To investigate sensitivity of diagnostic vitrectomy and vitreous biopsy for patients with vitritis of unknown aetiology. METHODS: Retrospective analysis of all vitrectomies and vitreous biopsies, performed at St Thomas' Hospital, London, UK, for vitritis between February 2001 and February 2019. Patients were identified using the VITREOR database and records were reviewed. Patients were categorised as infectious, non-infectious or masquerade based on final diagnosis. Sensitivity of both diagnostic pars plana vitrectomy (PPV) and vitreous cutter biopsy in each category was investigated. Furthermore, data on gender, age, and method of anaesthesia were also collected. RESULTS: In our cohort, 64 patients underwent PPV with a diagnostic sensitivity of 67% (43/64) overall and 60% (18/30), 56% (9/16) and 89% (16/18) for those with infectious, masquerade and non-infectious aetiologies, respectively. In comparison, 96 patients underwent a vitreous cutter biopsy with diagnostic sensitivity of 74% (71/96) overall and 71% (55/77), 67% (4/6) and 92% (12/13) for those with infectious, masquerade and non-infectious aetiologies, respectively. No statistically significant difference in sensitivity was identified between the vitrectomy and vitreous biopsy groups for either aetiology. Patients undergoing vitrectomy were noted to be older (p = 0.02) and more likely to undergo a general anaesthetic (p < 0.01). CONCLUSIONS: Herein we demonstrate similar diagnostic sensitivity of PPV and vitreous cutter biopsy in patients with vitritis of unknown aetiology.

Update in Molecular Testing for Intraocular Lymphoma.

The diagnosis of primary vitreoretinal lymphoma and central nervous system lymphoma is challenging. In cases with intraocular involvement, vitreous biopsy plays a pivotal role. Several diagnostic tests are employed to confirm a diagnosis and include cytologic evaluation, immunohistochemistry, flow cytometry, and cytokine analysis. The limitations of these conventional diagnostic tests stem from the often paucicellular nature of vitreous biopsy specimens and the fragility of malignant cells ex vivo. Several emerging molecular techniques show promise in improving the diagnostic yield of intraocular biopsy, possibly enabling more accurate and timely diagnoses. This article will review existing diagnostic modalities for intraocular lymphoma, with an emphasis on currently available molecular tests.

Vitreoretinal lymphoma following primary testicular lymphoma: Report of two cases and review of the literature.

PURPOSE: To report two cases of vitreoretinal lymphoma that developed following primary testicular lymphoma and review the literature. OBSERVATIONS: Two men, one age 66 and the other age 77, both with a history of diffuse large B-cell testicular lymphoma, diagnosed one and three years previously, respectively, presented with vitritis and yellow-white subretinal infiltrates. Diagnostic vitrectomy in both cases revealed diffuse large B-cell lymphoma. Systemic work up in both cases showed no evidence of disease relapse elsewhere. Each were treated with intravitreal methotrexate injections. CONCLUSIONS: Vitreoretinal lymphoma can occur following primary testicular lymphoma, and may mimic primary vitreoretinal lymphoma. Monitoring of patients with a history of testicular lymphoma with regular dilated fundus examinations should be considered.

Obtaining undiluted vitreous sample using small gauge pars plana vitrectomy and air infusion.

Vitreous biopsy is essential for establishing a definitive diagnosis and guiding potential therapy in patients with refractory vitritis and posterior uveitis, among other indications. Especially in cases where one must rule out intraocular lymphoma, obtaining a significant amount of undiluted vitreous increases diagnostic yield. We demonstrate our technique for obtaining a significant amount of undiluted vitreous using pars plana vitrectomy with air infusion.

Intraocular biopsy in uveitis.

An intraocular biopsy is performed for diagnostic, prognostic and investigational purposes. Biopsies help to confirm or exclude malignancies and differentiate inflammatory from infectious processes. Histopathological analysis is the final verdict in unresponsive uveitis, atypical inflammation, metastases and masquerade syndromes. Advances and refinement of techniques in cytopathology, immunohistochemistry, microbiological and molecular biologic study offer much more than just diagnosis. They provide prognosis based on cell characteristics and are helpful in planning treatment and intervention. Many biopsy procedures have evolved to provide more safety and minimise complications thus improving the quality of specimens or samples available for analysis. The type of biopsy and technique adopted varies based on the clinical suspicion, size and location of lesions. In uveitis, a working diagnosis of intraocular inflammation is made on clinical examination and laboratory investigations and ancillary tests. Malignancy and uveitis is interlinked and masquerade syndromes are among the commonest indications for biopsy and analysis of specimen. The various types of intraocular biopsies include aqueous tap, fine needle aspiration biopsy, vitreous biopsy, iris and ciliary body, and retinochoroidal biopsy. They will be reviewed in this article with respect to current perspective.

Comparison of microbiology and visual outcomes of patients undergoing small-gauge and 20-gauge vitrectomy for endophthalmitis.

BACKGROUND: The role of pars plana vitrectomy (PPV) for endophthalmitis has evolved over recent decades but the literature is lacking on comparisons between small-gauge and 20-gauge vitrectomy. OBJECTIVE: To evaluate evolving etiological and microbiological trends in patients undergoing vitrectomy for endophthalmitis and to compare culture-positive rates and visual outcomes between small-gauge (23- and 25-gauge) and 20-gauge instrumentation during vitrectomy for endophthalmitis. METHODS: Ten-year retrospective comparative case series and prospective laboratory in vitro testing. Tertiary care academic referral center. Patients who underwent PPV for endophthalmitis between 2003 and 2013. Vitreous biopsies were obtained in all cases. The effect of vitrectomy gauge (20-, 23-, and 25-gauge) and vitreous cutting rate (1,500 and 5,000 cuts per minute) on the viability of bacterial culture was evaluated in an in vitro prospective laboratory investigation. MAIN OUTCOME MEASURES: Comparison of etiology, microbiology culture-positive rates, and visual outcomes between small-gauge and 20-gauge instrumentation in patients undergoing PPV for infectious endophthalmitis. RESULTS: A total of 61 cases of vitrectomy for endophthalmitis were identified over a 10-year period; of these, 34 were treated with small-gauge (23- and 25-gauge) vitrectomy and 27 were treated with 20-gauge vitrectomy. In the small-gauge group, 12 cases (35.3%) yielded culture-positive results versus 20 cases (74.1%) with culture positivity in the 20-gauge cohort (P=0.002). The most common cause of endophthalmitis was cataract surgery and the most frequently identified organism was coagulase-negative Staphylococci in both groups. There was no significant difference in mean postoperative visual acuities between groups (P=0.33). Etiological trends indicate an increase in endophthalmitis due to intravitreal injection in the small-gauge group (n=9) compared to the 20-gauge group (n=3) (P=0.001). In vitro laboratory testing revealed no significant difference in rates of culture growth for different vitrectomy gauge sizes or vitreous cutting speeds. CONCLUSION AND RELEVANCE: Small-gauge vitrectomy for endophthalmitis yields final visual outcomes comparable to 20-gauge instrumentation. A significant difference in culture-positive rates was observed between small-gauge and 20-gauge instrumentation for vitrectomy in endophthalmitis; however, laboratory testing indicates this is not related to either vitreous gauge size or cutter speed. Intravitreal injections are emerging as a common etiology of vitrectomy for endophthalmitis.