Conjunctival Samples Research Articles

Prevalence of Leishmania infantum in felines from Heraklion region, Crete

Greece is recognized as an endemic region for various species of Leishmania, a genus of parasitic protozoa transmitted through the bites of infected female sandflies. This indicates that the country serves as a natural reservoir for this parasite, infecting both humans and animals. This study focuses on Heraklion, Crete, one of Greece's largest cities, where there is currently no available data on the prevalence of leishmaniosis in the feline population. While cases of canine leishmaniosis have been documented in various parts of the island, there have been no reported cases of feline leishmaniosis in Heraklion. This study primarily investigates the prevalence of leishmaniosis in cats as a means of detection in this species in Heraklion. To accomplish this, between 2018 – 2023, EDTA-blood samples and conjunctival swab samples were collected from a total of 253 feral cats. The cats were divided into two groups, depending on the location they lived (i.e., areas out of the city center or close to/in the city center): the initial group consisted of 112 cats, while the latter group comprised 141 cats. Both groups encompassed cats of various ages, habitats, and both genders. The samples were stored at -18 °C and analyzed with TaqMan real-time quantitative PCR for the presence of Leishmania infantum DNA.Out of the cat population included in the study, 15 were tested positive for FeL (5.93%). Despite the lower prevalence of leishmaniosis compared to dogs, it should be taken into account, especially for differential diagnosis. In conclusion, to the best of our knowledge, this is the first study that confirms the presence of FeL in felines in Heraklion.

Etiology and Outcomes of Acute Infectious Conjunctivitis in Children

To determine the causes of conjunctivitis and whether clinical presentations and outcomes differ by pathogen. This multicenter, case-control study enrolled 390 children (194 cases, 196 controls) whose conjunctival samples were tested for bacterial and viral pathogens. Caregivers completed surveys tracking symptoms, antibiotic use, school attendance, and adverse events. The outcomes analyzed included the prevalence of microorganisms detected by PCR in cases versus controls, symptoms, rate of resolution by day 5, school/childcare attendance, and parent-reported antibiotic-related adverse incidents. Most cases (148, 76%) and controls (112, 57%) had bacteria identified, although only detection of Haemophilus influenzae was associated with conjunctivitis (aOR 4.59, 95% CI 2.86, 7.37). Purulent discharge was associated with H. influenzae (aOR 2.47, 95% CI 1.23, 5.01) and occurred in 92 (77%) cases in which H. influenzae was detected and 39 (53%) in which H. influenzae was not detected. Improvement (186, 96%) and resolution (166, 86%) were observed by day 5 for most children and did not differ based on ophthalmic antibiotic use. Caregivers reported antibiotic-associated adverse events for 21 (20%) children with 8 (8%) requiring a medical visit. Only H. influenzae was significantly associated with conjunctivitis. Symptoms did not differ in children with or without bacteria detected by PCR. Independent of antibiotic use, most children experienced resolution by day 5, but parents reported adverse events in 20% of children treated with topical antibiotics, underscoring the importance of judicious prescribing.

The Occurrence of Chlamydia felis in Cats and Dogs in Hungary.

The World Health Organization (WHO) estimates that many human infections are zoonoses, creating a worldwide public health challenge. Among Chlamydia species, Chlamydia felis is the leading cause of conjunctivitis in cats and is a prominent zoonotic species. This study aimed to determine the occurrence and risk of chlamydiosis in cats and dogs in Szeged, Hungary, and surrounding areas. The total nucleic acids from conjunctival swab samples of symptomatic and asymptomatic animals were extracted using an automated nucleic acid extraction system. After that, DNA was amplified by pan-chlamydia PCR. Bacterial and fungal cultures were also performed to detect other microorganisms. Of the 93 animals, 32 (34.4%) were positive for pan-chlamydia PCR. The positivity rates were 33.3% (26/78) in cats and 40.0% (6/15) in dogs. Furthermore, the positivity rates were 37.2% (16/43) in the cat shelter, 42.4% (14/33) in the veterinary clinic, and 11.7% (2/17) in household pets. In total, 103 species were identified through culture-based examinations, including 97 (94.2%) bacterial and 6 fungal (5.8%) species. From both human and animal health perspectives, it is essential to have a detailed understanding of the circumstances of chlamydiosis, given the global impact of zoonotic diseases.

Predictive Value of Conjunctival Cytology in Bleb-dependent Glaucoma Surgery.

Metaplasia, chronic inflammation and subconjunctival fibrosis favor failure of bleb-dependent glaucoma surgery. The aim of the study is to identify the patients at a higher risk of post-surgical failure. Prospective, open study, performed in the Glaucoma Unit of the Hospital Universitario Virgen Macarena, from April to November 2021, with a minimum follow-up of one year. 38 eyes with ocular hypertension or chronic open-angle glaucoma were included. All patients underwent preoperative conjunctival sampling in the operating room, under topical or locoregional anesthesia. Sex, age, and laterality; number, type and mean time of preoperative drugs use; type of surgery performed; cytology results and degree of metaplasia; percentage of patients in whom the bleb was closed. Evaluation of potential correlation between bleb closure and any of the other variables. 20 women and 18 men participated, with a mean age of 67 years. The mean number of preoperative hypotensive drugs was 2.7.The mean time of use was 90,97 +/- 48,97 months. Most patients had normal cytology, 8% had inflammatory infiltrate and 21% had squamous metaplasia. When relating bleb failure and cytology, we saw that in those who failed surgery, more than half had cytological alterations. A multiple logistic regression was performed, in which we observed that there was statistically significant association (p = .02) between surgical closure and altered cytology. According to these results, preoperative conjunctival cytology can help predict those cases with a lower probability of surgical success.

Biomarker Detection and Validation for Corneal Involvement in Patients With Acute Infectious Conjunctivitis

Infectious conjunctivitis can lead to corneal involvement and result in ocular morbidity. The identification of biomarkers associated with corneal involvement has the potential to improve patient care. To identify biomarkers in patients with acute infectious conjunctivitis. This cross-sectional study took place from December 2016 to March 2024. Analyses were performed in 3 phases. First, logistic regression and machine learning algorithms were used to predict the probability of demonstrating corneal involvement in patients with presumed infectious conjunctivitis. Second, quantitative reverse transcription polymerase chain reaction (RT-qPCR) was used to confirm the most important biomarker gene identified by the algorithm. Third, the biomarker gene was validated in prospectively collected conjunctival samples of adult patients from 3 outpatient centers in Thailand and 1 in India. Patients with signs and symptoms of infectious conjunctivitis and onset within less than 14 days were eligible. Exclusion criteria were the inability to consent, presumed toxicity, or allergic conjunctivitis. Acute infectious conjunctivitis. The identification and validation of ocular surface gene expression associated with corneal findings on slitlamp examination. Thirteen genes exhibited a 1.5-log2 fold change in expression in patients with corneal involvement compared to patients without corneal involvement. Using the 13 genes to train and cross validate, logistic regression produced the highest mean area under the receiver operating characteristic curve (AUROC; 0.85; 95% CI, 0.84-0.86) for corneal involvement. The removal of apolipoprotein E (APOE) from the gene ensemble led to a decline in predictive performance of the logistic regression classifier (from mean AUROC 0.85 [95% CI, 0.84-0.86] to 0.74 [95% CI, 0.73-0.75]; adjusted P = .001 [Tukey test]). Orthogonal testing of APOE expression level with RT-qPCR showed that APOE expression was higher in patients with corneal involvement compared to patients without (median [IQR], 0.23 [0.04-0.47] vs 0.04 [0.02-0.06]; P = .004 [Mann-Whitney U test]). Using a Youden index of 0.23 Δ threshold cycle, APOE had a sensitivity of 56% (95% CI, 33-77) and a specificity of 88% (95% CI, 79-93) in 106 samples with conjunctivitis at Aravind, India (P < .001 [Fisher exact test]). When applied to a different patient population in Thailand, the same criteria could discriminate between disease states (58 samples; sensitivity, 47%; 95% CI, 30-64 and specificity, 93%; 95% CI, 77-99; P = .001 [Fisher exact test]). The results from this study suggest that the host conjunctival immune response can be meaningfully interrogated to identify biomarkers for ocular surface diseases.

Clinical insights and real-time PCR analysis of the first adeno-enterovirus conjunctivitis outbreak in India: A comprehensive cross-sectional study.

This cross-sectional study aimed to investigate the emergence of adeno-enterovirus conjunctivitis, with a focus on its clinical manifestations, prevalence, and implications within the context of acute epidemic conjunctivitis. Real-time PCR analysis was employed to precisely identify the viral strain in conjunctival swab samples. A total of 80 patients were tested, and specific clinical symptoms were assessed. The study revealed the first documented case in India and the second worldwide of adeno-enterovirus conjunctivitis. The prevalence among the tested patients was significant, with 15% testing positive. Those infected with adeno-enterovirus exhibited shorter illnesses (≤3 days). Notably, clinical symptoms such as subconjunctival hemorrhage, chemosis, swelling, photophobia, and epiphora were significantly linked to this viral variant. Multivariable logistic regression analysis highlighted swelling and photophobia as robust predictors for identifying adeno-enterovirus conjunctivitis cases. The findings underscore the urgent need to recognize adeno-enterovirus conjunctivitis as a potential threat, capable of causing significant ocular morbidity. Swelling and photophobia emerged as robust predictors for identifying cases, emphasizing the importance of vigilant monitoring and comprehension of this viral variant. Proactive measures are essential to address its clinical implications and mitigate its impact on affected populations. Ongoing research is crucial to unravel the complete spectrum of adeno-enterovirus conjunctivitis and guide effective management strategies.

SARS-CoV-2 positivity rates in tear secretions of inpatient COVID-positive individuals in an Indian tertiary care setting.

In the third wave of the coronavirus disease 2019 (COVID-19) pandemic, despite progressive vaccination coverage, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was rapidly spreading and the possibility for ocular transmission of COVID-19 was most relevant. This study aimed to determine the presence of SARS-CoV-2 RNA in tears as detected by real-time polymerase chain reaction (RT-PCR), among individuals who tested COVID-positive during the third wave of the COVID pandemic. Among 90 patients admitted with mild to severe COVID-19 infection (ranging from asymptomatic to severe symptoms), separate conjunctival samples were obtained from both eyes without topical anesthesia taking precautions to avoid contamination. Inferior fornices of the eyes were swept slowly with sterile Dacron swabs for 10 s or until visibly wet. They were inoculated directly in a viral lysis buffer containing carrier RNA and transported to the Virology Laboratory for the detection of SARS-CoV-2 RNA by RT-PCR. Conjunctival swabs were declared positive based on cycle threshold values of <37. Seven of 180 conjunctival swabs were positive (3.9%). Five patients had unilateral positive and one patient had bilateral positive conjunctival swabs. Thirty-six (40%) patients had their conjunctival swabs taken on the same day of detection of the first positive nasopharyngeal swab, while the rest (54) had their conjunctival swabs taken up to 3 days later. Overall, 20 (22.2%) patients were symptomatic for COVID-19. Considering the contagious nature of SARS-CoV-2 in the third wave, there is a possibility that ocular surface fluid might have been a contributing factor along with other modes of transmission of COVID-19.

Single-cell sequencing depicts tumor architecture and empowers clinical decision in metastatic conjunctival melanoma

Conjunctival melanoma (CoM) is a potentially devastating tumor that can lead to distant metastasis. Despite various therapeutic strategies for distant metastatic CoM, the clinical outcomes remain unfavorable. Herein, we performed single-cell RNA sequencing (scRNA-seq) of 47,017 cells obtained from normal conjunctival samples (n = 3) and conjunctival melanomas (n = 7). Notably, we noticed a higher abundance of cancer-associated fibroblasts (CAFs) in tumor microenvironment (TME), correlated with enhanced angiogenic capacity and increased VEGFR expression in distal metastatic CoM. Additionally, we observed a significant decrease in the proportion of total CD8+ T cells and an increase in the proportion of naive CD8+ T cells, contributing to a relatively quiescent immunological environment in distal metastatic CoM. These findings were confirmed through the analyses of 70,303 single-cell transcriptomes of 7 individual CoM samples, as well as spatially resolved proteomes of an additional 10 samples of CoMs. Due to the increase of VEGFR-mediated angiogenesis and a less active T cell environment in distal metastatic CoMs, a clinical trial (ChiCTR2100045061) has been initiated to evaluate the efficacy of VEGFR blockade in combination with anti-PD1 therapy for patients with distant metastatic CoM, showing promising tumor-inhibitory effects. In conclusion, our study uncovered the landscape and heterogeneity of the TME during CoM tumorigenesis and progression, empowering clinical decisions in the management of distal metastatic CoM. To our knowledge, this is the initial exploration to translate scRNA-seq analysis to a clinical trial dealing with cancer, providing a novel concept by accommodating scRNA-seq data in cancer therapy.

Next-Generation Sequencing of the Human Aqueous Humour Microbiome.

The microbiome of the ocular surface has been characterised, but only limited information is available on a possible silent intraocular microbial colonisation in normal eyes. Therefore, we performed next-generation sequencing (NGS) of 16S rDNA genes in the aqueous humour. The aqueous humour was sampled from three patients during cataract surgery. Air swabs, conjunctival swabs from patients as well as from healthy donors served as controls. Following DNA extraction, the V3 and V4 hypervariable regions of the 16S rDNA gene were amplified and sequenced followed by denoising. The resulting Amplicon Sequence Variants were matched to a subset of the Ribosomal Database Project 16S database. The deduced bacterial community was then statistically analysed. The DNA content in all samples was low (0-1.49 ng/µL) but sufficient for analysis. The main phyla in the samples were Acinetobacteria (48%), Proteobacteria (26%), Firmicutes (14%), Acidobacteria (8%), and Bacteroidetes (2%). Patients' conjunctival control samples and anterior chamber fluid showed similar patterns of bacterial species containing many waterborne species. Non-disinfected samples showed a different bacterial spectrum than the air swab samples. The data confirm the existence of an ocular surface microbiome. Meanwhile, a distinct intraocular microbiome was not discernible from the background, suggesting the absence of an intraocular microbiome in normal eyes.

Molecular characterization of coxsackievirus A24 variants isolated from an outbreak of acute hemorrhagic conjunctivitis

PurposeAcute Hemorrhagic conjunctivitis (AHC) is associated with CVA24v. Recently there was a severe outbreak of conjunctivitis in months of July and August 2023 in India. This study emphasizes the identification of the distinct mutations in the CVA24v strains, which were isolated during the AHC outbreak and could have potentially played a role in the high transmission of AHC in India during the 2023 outbreak. MethodsA total of 71 conjunctivitis patients aged 1–75 years comprising 47 males and 24 females who attended Ophthalmology department of a tertiary care hospital of easternIndia were studied.RNA was extracted from all conjunctival swab samples and converted into cDNA. Subsequently, the viral 5′ UTR was amplified and the PCR positive samples were subjected to sequencing. The newly isolated viral 5′ UTR sequences were aligned with other worldwide sequences using the Clustal W tool to conduct mutational analysis. A phylogenetic tree was built using the MEGA software for viral genotype identification. ResultsAll of the current outbreak strains belonged to genotype IV of CVA24v. The present outbreak strains formed a distinct clade in the phylogenetic tree and were different from previously reported Indian strains. Two persistent mutations, specifically in domain IV (T213C) and domain V (C475T), were exclusively detected within the internal ribosome entry site (IRES) of the 5′ UTR of the current strains causing the outbreak. These two alterations have previously been shown to impact the virulence of another enterovirus (CV B3), but they have not been described in CVA24v until now. ConclusionFinding of the present study highlights the possibility and the significance of the aforementioned two mutations in enhancing the transmissibility of the newer CVA24v strains. Hence, these two distinct mutations should be investigated further for developing antiviral therapies to combat future AHC outbreaks associated with CVA24v.

Phospholipid analyses of rabbit ocular surface tissues

The tissues of the integument covering the ocular surface comprise a mucus membrane functioning as a protective physical barrier and has the ability to mount a defensive inflammatory response. Since lipid metabolism has a role in both of these functions, we studied normal membrane phospholipids (PL) of the cornea and bulbar conjunctiva to (1) determine baseline PL profiles of these tissues, (2) compare and contrast these individual PL metabolite profiles as well as groups of metabolites, and (3) describe pathway-specific metabolic interrelations among these tissues. Corneal and conjunctival tissue samples were isolated from rabbit eyes (n = 30) and extracted with chloroform-methanol using a modified Folch procedure. 31P nuclear magnetic resonance spectroscopy was used to qualitatively and quantitatively measure tissue PL profiles. The cornea and conjunctiva, respectively, have the following PL composition (mole % of total detected phospholipid): phosphatidylglycerol (PG) –, 0.4; lysophosphatidylethanolamine 1.2, –; phosphatidic acid –, 0.4; diPG (cardiolipin) 2.1, 3.5; unknown PL at the chemical shift of 0.13 δ 1.5, 0.9; ethanolamine plasmalogen 11.2, 13.0; phosphatidylethanolamine 11.5, 12.8; phosphatidylserine 8.9, 10.1; sphingomyelin 10.2, 10.7; lysophosphatidylcholine 0.9, 1.4; phosphatidylinositol 5.3, 5.3; phosphatidylcholine (PC) plasmalogen or alkylacylPC 2.2, 1.9; PC 45.1, 40.0. In addition, 28 PL metabolic indices were calculated from these data, which permitted pathway-specific lipid analyses. This study (1) establishes PL profiles of the two ocular tissues of the integument that cover the surface of the eye, (2) compares and contrasts indices comprised of ratios and combinations of PL, and (3) describes pathway-specific metabolic interrelations among these tissues to serve as baselines for studies involving the distribution of tissue phospholipids.

Expression of CD44, PCNA and E-cadherin in pterygium tissues.

Pterygium is a common ocular surface disease defined by fibrovascular conjunctival growth extending onto the cornea. However, its pathogenesis remains unclear. This study aimed to determine the role of CD44, proliferating cell nuclear antigen (PCNA), and E-cadherin in pterygium formation and recurrence. Sixty patients with pterygium participated in the study, and we collected conjunctival samples from 30 patients to form a control group. CD44, PCNA, and E-cadherin expressions in surgically excised pterygium were compared with tissue samples from the control group. We observed that the percentages of CD44 and PCNA were statistically higher in the primary pterygium group and recurrent pterygium group than in the control group (P < 0.001 and P < 0.001, respectively). Conversely, E-cadherin values were statistically higher in the control group than in the primary and recurrent pterygium groups (P = 0.013 and P < 0.001, respectively). Cell proliferation and cell adhesion factors may play important roles in the pathogenesis of pterygium.

Feline leishmaniasis in an animal shelter in northeastern Brazil: Clinical aspects, coinfections, molecular detection, and serological study of a new recombinant protein

Infection and clinical cases of leishmaniasis caused by Leishmania infantum in cats have been increasingly reported in several countries, including Brazil. In this study, we used an enzyme-linked immunosorbent assay (ELISA) and an immunochromatographic test (ICT) based on a recombinant antigen (rKDDR-plus) to detect anti-Leishmania antibodies in cats from an animal shelter in northeastern Brazil. We compared the results with an ELISA using L. infantum crude antigen (ELISA-CA). We also investigated the presence of Leishmania DNA in blood or ocular conjunctival samples as well as the association between Leishmania PCR positivity and serological positivity to feline immunodeficiency virus (FIV), feline leukemia virus (FeLV) and Toxoplasma gondii. Concerning serological assays, a higher positivity was detected using the ICT-rKDDR-plus (7.5%; 7/93) as compared to ELISA-rKDDR-plus (5.4%; 5/93) and ELISA-CA (4.3%; 4/93). Upon PCR testing, 52.7% (49/93) of the ocular conjunctival swabs and 48.3% (44/91) of the blood samples were positive. Together, PCR and serological testing revealed overall positivities of 73.1% (68/93) and 12.9% (12/93), respectively. Among PCR-positive samples, 45.5% (31/68) showed co-infection with FIV, 17.6% (12/68) with FeLV, and 82.3% (56/68) with T. gondii. More than half of the PCR-positive cats showed at least one clinical sign suggestive of leishmaniasis (58.8%; 40/68) and dermatological signs were the most frequent ones (45.5%; 31/68). Both tests employing the recombinant antigen rKDDR-plus (i.e., ICT-rKDDR-plus and ELISA-rKDDR-plus) detected more positive cats than the ELISA-CA but presented low overall accuracy. PCR testing using either blood or ocular conjunctival samples detected much more positive cats than serological tests.

Dry eye symptoms are prevalent in moderate-severe COVID-19, while SARS-COV-2 presence is higher in mild COVID-19: Possible ocular transmission risk of COVID-19

PurposeTo evaluate the correlation between dry eye symptoms and coronavirus disease 2019 (COVID-19) infection and to assess the real-time reverse transcription–polymerase chain reaction (RT‒PCR) of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) from the conjunctival swab. MethodsA prospective observational case series study was conducted of all suspected and confirmed COVID-19 patients from Dr. Cipto Mangunkusumo Hospital (RSCM) and the Universitas Indonesia Hospital (RSUI). On the first day of the visit (day 0), systemic clinical symptoms and naso-oropharyngeal (NO) RT‒PCR results will classify all subjects as non-, suspected, or confirmed (mild, moderate, and severe) COVID-19. In all patients, we determined the dry eye symptoms based on the Ocular Surface Disease Index (OSDI) and followed up 7(day 7) and 14 days (day 14) after the first visit. When it was technically possible, we also examined the objective dry eye measurements: tear meniscus height (TMH), noninvasive Keratograph® break-up time (NIKBUT), and ocular redness. Additionally, we took conjunctival swab samples for SARS-CoV-2 RT-PCR in all patients. ResultsThe OSDI scores for 157 patients decreased across days 0, 7, and 14 (median (interquartile range): 2.3 (0–8), 0 (0–3), and 0 (0-0), p value < 0.0001 (D0 vs D14). The moderate-severe COVID-19 group had a higher OSDI score than the other groups at median D0 (15.6 vs 0–2.3), p value < 0.0001 and this pattern was consistently seen at follow-up D7 and D14. However, dry eye complaints were not correlated with the three objective dry eye measurements in mild-moderate COVID-19 patients. NO RT‒PCR results were positive in 32 (20.4%) patients, namely, 13 and 19 moderate-severe and mild COVID-19 patients, respectively. Positive RT‒PCR results were observed in 7/157 (4.5%) conjunctival swab samples from 1 in non-COVID-19 group and 6 in mild group. ConclusionIn the early phase of infection, COVID-19 patients experience dry eye symptoms, which have no correlation with objective dry eye measurements. SARS-CoV-2 in conjunctival swab samples can be detected in patients with normal-to-mild COVID-19, which shows the risk of ocular transmission.

Dupilumab‐associated ocular surface disease or atopic keratoconjunctivitis not improved by dupilumab? Upadacitinib may clarify the dilemma: A case report

AbstractDupilumab‐associated ocular surface disease is a common clinical sign appearing in patients with atopic dermatitis (AD) just few months after dupilumab treatment start, developing in about 25% of patients. Atopic keratoconjunctivitis (AKC) is a well‐identified clinical entity, defined as a chronic inflammatory disease of eye that affects 25%–40% of patients with AD. Most clinical signs of ocular involvement in AD patients treated with dupilumab overlaps the AKC symptoms and signs. We supposed that Dupilumab‐associated ocular surface disease and AKC represent the same disease but differently called by dermatologists and ophthalmologists. AKC‐like disease may develop during dupilumab therapy as a consequence of alternative cytokines pathway activation (e.g. IL33) secondary to IL‐4/13 pathway block. The novel upadacitinib drug may bypass ILs pathway through Janus Kinases selective inhibition, avoiding positive or negative ILs feedback at the ocular surface level. In this case report, molecular analysis on conjunctival samples showed a lower ocular surface inflammation (lower expression of HLADR) although higher levels of IL4 and IL13 in a patient with AD and AKC during upadacitinib therapy, compared to prior dupilumab treatment. Target therapies in patients suffering from AD may prevent ocular and dermatological comorbidities improving quality of life before quality of skin and vision.

Analysis of the mucosal chemokines CCL28, CXCL14, and CXCL17 in dry eye disease: An in vitro and clinical investigation

Mucosal chemokines have antimicrobial properties and play an important role in mucosal immunity. However, little is known about their expression on the ocular surface. This study aimed to analyze the expression of the mucosal chemokines CCL28, CXCL14 and CXCL17 in corneal and conjunctival epithelial cells under in vitro dry eye (DE) conditions, and in conjunctival samples from healthy subjects and DE patients. Human corneal epithelial cells (HCE) and immortalized human conjunctival epithelial cells (IM-HConEpiC) were incubated under hyperosmolar (400–500 mOsM) or inflammatory (TNF-α 25 ng/mL) conditions for 6 h and 24 h to measure CCL28, CXCL14, and CXCL17 gene expression by RT-PCR and their secretion by immunobead-based analysis (CCL28, CXCL14) and ELISA (CXCL17). Additionally, twenty-seven DE patients and 13 healthy subjects were included in this study. DE-related questionnaires (OSDI, mSIDEQ and NRS) evaluated symptomatology. Ocular surface integrity was assessed using vital staining. Tactile sensitivity was measured with Cochet-Bonnet esthesiometer, and mechanic and thermal (heat and cold) sensitivity using Belmonte's non-contact esthesiometer. Subbasal nerve plexus and dendritic cell density were analyzed by in vivo confocal microscopy. Conjunctival cells from participants were collected by impression cytology to measure mucosal chemokines gene expression by RT-PCR. Our results showed that HCE and IM-HConEpiC cells increased CCL28, CXCL14, and CXCL17 secretion under hyperosmolar conditions. The gene expression of CCL28 was significantly upregulated in conjunctival samples from DE patients. CCL28 expression correlated positively with symptomatology, corneal staining, heat sensitivity threshold, and dendritic cell density. CXCL14 expression correlated positively with age, ocular pain, conjunctival staining, tactile sensitivity, and image reflectivity. CXCL17 expression correlated positively with corneal staining. These results suggest that corneal and conjunctival epithelial cells could be a source of CCL28, CXCL14, and CXCL17 on the ocular surface and that CCL28 might be involved in DE pathogenesis.

An outbreak of acute hemorrhagic conjunctivitis due to Coxsackievirus A24 in a residential school, Naharlagun, Arunachal Pradesh: July 2023

PurposeAn acute conjunctivitis outbreak was investigated at a residential school in Naharlagun, Arunachal Pradesh, Northeast India, in July 2023. We aimed to identify the etiological agent and assess any complications in follow-up cases. MethodsWe used a structured questionnaire to record clinical findings and followed up with cases one-month post-conjunctivitis. Sixty-one cases were examined and eight conjunctival and oropharyngeal swab samples were collected after obtaining informed consent from guardians/school authorities. We screened for 33 viral and bacterial pathogens using an IVD-approved Real-time PCR assay. Further, the samples were subjected to nucleic acid sequencing. ResultsAmong 465 screened students and staff, 80 individuals (approximately 17.2%) showed acute hemorrhagic conjunctivitis symptoms among which 61 cases were available for clinical examination. We identified the Enterovirus responsible by targeted sequencing using next-generation sequencing. The etiological agent was found to be Coxsackievirus A24, a member of Enterovirus C, in seven out of eight samples subjected to sequencing. Common symptoms included conjunctival hyperemia and foreign body sensation (100%), bilateral eye involvement (73.8%), eye pain (70%), watery discharge (49.2%), and eyelid swelling (38%). Only 6.5% had purulent discharge. Most cases resolved within 5-6 days, with only 9.8% reporting abdominal symptoms post-conjunctivitis. No serious complications occurred within one month. Throat swabs aided in diagnosing enterovirus infections alongside eye swabs. ConclusionsThe outbreak of acute conjunctivitis was caused by Coxsackievirus A24, a member of Enterovirus C. Cases resolved spontaneously within 6-7 days, with no severe complications. Collecting oropharyngeal swabs alongside conjunctival swabs could improve enteroviral conjunctivitis diagnosis.

Molecular detection and characterization of Acanthamoeba infection in dogs and its association with keratitis in Korea.

Acanthamoeba infection is associated with keratitis in humans; however, its association with keratitis in dogs remains unclear. To investigate this possibility, we collected 171 conjunctival swab samples from dogs with eye-related diseases (65 with keratitis and 106 without keratitis) at Chungbuk National University Veterinary Teaching Hospital, Korea, from August 2021 to September 2022. Polymerase chain reaction identified 9 samples (5.3%) as Acanthamoeba positive; of these, 3 were from dogs with keratitis (4.6%) and 6 were from dogs without keratitis (5.7%). Our results indicated no significant association between Acanthamoeba infection and keratitis, season, sex, or age. All Acanthamoeba organisms found in this study had the genotype T4, according to 18S ribosomal RNA analysis. Acanthamoeba infection in dogs might have only a limited association with keratitis.

Comparison of conjunctival swabs and blood samples in dogs for molecular and serological detection of Leishmania infantum infection in Colombia.

Surveillance of canine leishmaniasis in Colombia is restricted to the appearance of visceral leishmaniasis cases in humans, and is mainly performed by serological tests. This requires blood sampling by veterinarians or technicians according to Colombian laws. The aim of this study was to evaluate the utility of conjunctival swabs in the molecular detection of Leishmania in dogs from the municipality of Ovejas, Sucre. The present study was cross-sectional and descriptive. The collection source of samples and information was primary. Blood samples and conjunctival swabs from 121 dogs were analysed by PCR-ITS1 to detect Leishmania spp. Positive samples were used to amplify a conserved region of the Leishmania infantum kinetoplast minicircle. Performance of both sample types was calculated by proportion of positive samples of each type and the degree of agreement between them was determined by Cohen's kappa (κ) agreement index. Leishmania infection was detected in 17.4% (21/121) of blood samples and in 16.5% (20/121) of conjunctival swabs. In total, 28.1% (34/121) of the canines were infected, of which 11.8% (4/34) were infected with L. infantum in the conjunctival swabs and 5.9 % (2/34) in the blood samples. The agreement between blood and conjunctiva was medium (κ = 0.207) by PCR-ITS1 amplification. The use of conjunctival swab as a non-invasive sample could be used as an alternative method for surveillance of canine leishmaniasis.

Cycle threshold values of SARS-CoV-2 RNA in conjunctival swabs and nasopharyngeal secretions: a comparative study from a tertiary care center in India.

To detect the viral RNA load of SARS-CoV-2 in conjunctival swabs of COVID-19 patients, and compare with nasopharyngeal swabs. Conjunctival swabs of COVID-19 patients (with PCR positive nasopharyngeal swabs) were subjected to quantitative reverse transcription-polymerase chain reaction (RT-PCR) for detection of SARS-CoV-2 RNA. The cycle threshold (Ct) values of Open Reading Frame 1 (ORF 1 Ab gene) and nucleoprotein (N gene) PCRs were used to assess the viral RNA load, and compare them with the baseline values of nasopharyngeal swabs. Of 93 patients, 17 (18.27%) demonstrated SARS-CoV-2 RNA in conjunctival swabs. Baseline nasopharyngeal swabs were collected at a median of 2days; while, the conjunctival swabs were collected at median 7days, from onset of illness (p < 0.001). Despite a significant delay in conjunctival swab collection than nasopharyngeal swabs, the Ct values (ORF or N gene PCRs) were comparable between nasopharyngeal swab and conjunctival swab samples. Subsequently, during the recovery period, in four of these 17 patients (with conjunctival swab positivity), when the second nasopharyngeal swab was 'negative', the conjunctival swab was 'positive'. The conjunctival swabs demonstrated SARS-CoV-2 RNA in 17 (18.27%) of 93 COVID-19 patients. Our results may suggest a delayed or a prolonged shedding of the virus/viral RNA on the ocular surface than in nasopharyngeal mucosa.