H9N2 avian influenza virus-induced conjunctivitis model for vaccine efficacy testing.

Clinical signs such as respiratory signs, egg drop, and mortality have been reported in field cases of low pathogenic avian influenza by H9N2 avian influenza virus (AIV) but have rarely been reproduced by the virus alone. Thus, virus reisolation rates and titers in tissues were measured for vaccine efficacy testing. In the present study, we established a clinical sign-based vaccine efficacy test by reproduction of highly frequent conjunctivitis (77.8%-90%) via binocular instillation of an H9N2 virus (01310) strain, 1 x 10(6) EID50/10 microl for each eye). Specific-pathogen-free chickens were assigned to vaccine and control groups, and the vaccine group was inoculated intramuscularly with a commercial H9N2 inactivated oil emulsion vaccine. The chickens were challenged by 01310 via binocular instillation at 2 and 4 wk postvaccination (WPV). The positive rates of conjunctivitis and virus reisolation were significantly different between the vaccine and control groups (conjunctivitis at 2 WPV, 0% vs. 77.8%, and at 4 WPV, 0% vs. 80%). Vaccine antibody was detected in tears as well as in serum samples of the vaccine group before challenge. The conjunctivitis model may be useful for efficacy testing of AI vaccine due to a clinical symptom-based read of results, but further efficacy testings with different types, doses of AI vaccines, and challenge viruses will be required to complete the evaluation of our model.

DP2 (CRTh2) antagonism reduces ocular inflammation induced by allergen challenge and respiratory syncytial virus.

BACKGROUND: Allergic conjunctivitis is characterized by itchy, watery and swollen eyes which occur in response to exposure to seasonal or environmental allergens. The early phase reaction of allergic conjunctivitis is primarily mediated by mast cell degranulation while the late phase reaction is driven by Th2 cells and eosinophils. Prostaglandin D(2) (PGD(2)), released from mast cells, is present in allergic conjunctival tears and may elicit classical allergic responses via interaction with the high-affinity DP2 receptor (chemoattractant receptor-homologous molecule expressed on Th2 cells, CRTh2). Furthermore, antagonism of this receptor is well known to inhibit eosinophil chemotaxis, basophil activation and Th2 cytokine production. PGD(2), therefore, may be involved in both early and late phase reactions in response to allergen challenge. METHODS: Thus, we explored whether our novel and selective DP2 antagonist AM156 would be efficacious in animal models of allergic conjunctivitis. Furthermore, as respiratory syncytial virus (RSV) has been implicated in the pathogenesis of allergic conjunctivitis, we examined the effects of DP2 antagonism in a murine model of RSV ocular infection. RESULTS: Utilizing a guinea pig ovalbumin model and a murine ragweed model we demonstrated that AM156 reduces redness, discharge and swelling in response to allergen challenge. These effects were equal to or greater than those of current clinical treatment options for allergic conjunctivitis including topical corticosteroids and a dual-mechanism antihistamine and decongestant. AM156 significantly reduced RSV-induced ocular inflammation and IL-4 production. CONCLUSION: These results suggest that a topical DP2 antagonist such as AM156 may represent a novel therapeutic for allergic conjunctivitis.

Experimental primary ocular canine herpesvirus-1 infection in adult dogs.

OBJECTIVE-To characterize clinical ocular disease, viral shedding, and serologic response associated with primary canine herpesvirus-1 (CHV-1) ocular infection in naïve adult dogs. ANIMALS-12 specific pathogen-free adult Beagles. PROCEDURES-Dogs were topically inoculated in the right eye with CHV-1 (infection group; n = 8) or virus-free medium (control group; 4). Dogs were inoculated with or without corneal microtrephination and subconjunctivally administered corticosteroids. Conjunctiva, buffy coat, and serum samples for real-time PCR assay, virus isolation, and serum neutralization (SN) antibody titers were collected until postinfection day (PID) 224, and general physical and ophthalmologic examinations were performed. RESULTS-Dogs in the infection group developed bilateral, mild to moderate conjunctivitis that reached maximal intensity on PIDs 7 to 10. Ocular viral shedding was detected in all dogs in the infection group between PIDs 3 and 10. Infected dogs developed CHV-1 SN antibody titers, beginning at PID 7 and peaking on PID 21. All buffy coat PCR assay results were negative. Corneal microtrephination and subconjunctival corticosteroid administration did not significantly affect clinical disease or viral shedding. Following recovery from primary infection, dogs remained clinically normal, did not shed virus, and had slowly decreasing SN antibody titers. Dogs in the control group did not develop conjunctivitis, shed virus, or develop CHV-1 SN antibody titers. CONCLUSIONS AND CLINICAL RELEVANCE-Primary ocular infection of adult dogs with CHV-1 was associated with self-limiting conjunctivitis and ocular viral shedding, which was evident in the absence of clinically detectable keratitis or systemic disease. Features of this infection resembled herpes simplex virus primary ocular infection in humans.

[Chronic persistent infections of anterior segment of eye: clinico-laboratory aspects].

AIM: Improvement of therapy of chronic ophthalmic infectious diseases during assessment of functioning of different arms of immune system. MATERIALS AND METHODS: Three hundred and fifty patients with chronic red-eye syndrome were tested by immunofluorescence assay on the presence of antigens of herpesviruses, adenoviruses and Chlamydia in samples from conjunctiva. Expression of 11 cytokines' genes was measured in peripheral blood mononuclear cells by reverse transcription polymerase chain reaction. Production of IFN-alpha and IFN-gamma, levels of serum and spontaneously produced interferon as well as level of susceptibility to the range of immunomodulating preparations were measured during study of interferon status in whole blood cells. Study of parameters of cytokine, interferon and immune statuses was performed in 70 patients. Counts of T- and B-lymphocytes, T-helpers, NK-cells as well as level of circulating immune complexes were measured during study of immune status. RESULTS: Antigens of herpes simplex virus and adenoviruses were detected in samples from conjunctiva in 27% (95 persons) and 36% (126 persons) of patients respectively. Enhanced level of expression of several cytokines (IL-2, IL-4) in studied patients compared with healthy volunteers was observed. Expression levels of IL-12 and TNF-alpha mRNAs were, in opposite, in 2 - 3 times lower. Disorder of IFN-alpha and IFN-gamma synthesis on post-transciption level was observed in 60 - 90% of patients. Decrease of absolute numbers of total T-lymphocytes and T-helpers as well as increase of absolute number of NK-cells was noted in 20%, 25%, and 27% of patients respectively. CONCLUSION: Assignment of individually oriented antiviral, antibacterial and immunomodulating therapy allowed to mitigate intensity of clinical symptoms in 30 -60% of patients with chronic persistent infections of anterior segment of eye.

Serological evidence of Zika virus infection in febrile patients at Greater Accra Regional Hospital, Accra Ghana.

OBJECTIVE: Increase in the evidence of global occurrence of Zika viral infection suggests that in Africa the circulation of the virus which causes 80% of asymptomatic infection could be undetected and/or overlooked. We sought to serologically detect Zika virus infection in febrile patients at Greater Accra Regional Hospital, Ghana. RESULTS: Of the 160 patient serum samples analyzed, 33 were found to have antibodies against Zika virus infection. Among the sero-positives 30 (91%) of the cases were anti-Zika virus IgM with the 21-30-year age group recording the highest number of 8 (26%) and 2 (7%) cases being the least for the 61 years and above age group. All sero-positive febrile patients developed at least one symptom consistent with Zika virus infection: 33 (100%) fever, 25 (76%) muscle pain, 24 (73%) joint pain, and conjunctivitis 2 (6%). Digestive symptoms recorded include 16 (49%) nausea, 12 (36%) vomiting and diarrhea 18 (55%). In addition, 28 (85%) loss of appetite, 14 (75%) rapid respiration and chest pain 15 (42%) were reported by seropositive febrile patients. Our data indicates exposure to Zika virus which suggests the possible circulation of the virus among febrile patients in Ghana with a sero-prevalence rate of 20.6%.

Primary conjunctival herpetic geographic ulcer in an immunocompetent patient.

Herpes simplex virus ocular infections have been recognized as a major cause of corneal blindness in the developed world. The major signs of a primary infection are limited to the lids, conjunctiva, and cornea. Presence of a conjunctival dendrite without corneal involvement has been reported. Although conjunctival ulceration has been associated with recurrent attacks, it has not been documented as a manifestation of primary herpetic infection. We report a case of primary herpetic infection with geographic conjunctival ulcer with multiple corneal dendrites.

Allergy and infections: long-term improvement of vernal keratoconjunctivitis following viral conjunctivitis.

PURPOSE: Vernal keratoconjunctivitis (VKC) is a severe, chronic allergic inflammatory disease of the ocular surface poorly responsive to antiallergic treatments and possibly leading to permanent visual impairment. VKC, because of mast cell, eosinophil, and Th2-type inflammation, polyclonal IgE activation, and tissue remodeling, is considered to be a typical Th2- driven disease. Viral infection stimulates a Th1 type immune response, potentially attenuating allergen-induced inflammation. The purpose of this report is to describe the effect of viral keratoconjunctivitis in a patient with VKC. METHODS: The authors report on a patient with a severe form of VKC, poorly responsive to antiallergic treatments, who developed a viral keratoconjunctivitis. Signs, symptoms, and cytologic findings were recorded during the 5-year follow-up period. RESULTS: The authors observed a prompt and permanent improvement of signs and symptoms of the allergic condition after the viral infection. Conjunctival scraping confirms that the inhibition of the eosinophilic inflammation lasts at least for 5 years. CONCLUSIONS: In this case, the viral infection seemed to induce a clinical recovery of allergic disease, suggesting that an immune deviation induced by Th1-polarizing agents may revert an ongoing Th2 inflammation.

Dynamic change in natural killer cell type in the human ocular mucosa in situ as means of immune evasion by adenovirus infection.

The most severe form of virus-induced inflammation at the ocular surface is epidemic keratoconjunctivitis (EKC), often caused by group D human adenoviruses (HAdVs). We investigated the dynamics and mechanisms of changes in natural killer (NK) cell types in the human ocular mucosal surface in situ over the course of infection. In the acute phase of infection, the mature CD56(dim)NK cells that comprise a major subpopulation in the normal human conjunctiva are replaced by CD56(bright)NK cells recruited to the ocular surface by chemokines produced by the infected epithelium, and NKG2A-expressing CD56(dim) and CD56(bright) NK cells become the major subpopulations in severe inflammation. These NK cells attracted to the mucosal surface are however incapable of mounting a strong antiviral response because of upregulation of the inhibitory ligand human leukocyte antigen-E (HLA-E) on infected epithelium. Furthermore, group D HAdVs downregulate ligands for activating NK cell receptors, thus rendering even the mature NKG2A(-)NK cells unresponsive, an immune-escape mechanism distinct from other adenoviruses. Our findings imply that the EKC-causing group D HAdVs utilize these multiple pathways to inhibit antiviral NK cell responses in the initial stages of the infection.

Virological and molecular biological investigations into equine herpes virus type 2 (EHV-2) experimental infections.

Two 18-month-old naturally reared ponies were used to investigate the pathogenicity of EHV-2. After dexamethasone treatment, pony 1 was inoculated intranasally with EHV-2 strain T16, which has been isolated from a foal with keratoconjunctivitis superficialis and pony 2 was similarly inoculated with strain LK4 which was originally isolated from a horse with upper respiratory tract disease. Following virus inoculation, pyrexia was not detected in either pony but both developed conjunctivitis, lymphadenopathy, and coughing. EHV-2 was detected in nasal mucus samples up to day 12 post infection (p.i.), in eye swabs up to day 10 p.i., and in buffy coat cells throughout the investigation in both animals. EHV-2-specific antibody titres were raised significantly 18 days p.i. Following the administration of dexamethasone, 3 months p.i., infectious virus was again detected in nasal mucus and conjunctival swabs from both ponies for 7 days. The tissue distribution of EHV-2 genome was studied post mortem, by means of a nested PCR. EHV-2 was detected in lymphoid tissues, lung, conjunctiva, trigeminal ganglia and olfactory lobes of pony 2, whereas in pony 1 only the conjunctiva of the left eye was PCR positive.

Immunoassay of tear lysozyme in acute adenovirus conjunctivitis.

The tear lysozyme levels were measured by immunoassay in 92 healthy subjects and 98 patients with acute adenovirus conjunctivitis. They were found to be significantly decreased during the acute phase of the disease. The extent of this decline in the tear lysozyme level was correlated with increased severity of disease. There was no significant difference in the tear lysozyme level in viral isolation-positive and isolation-negative patients. The tear lysozyme level showed return to normal levels with clinical improvement.

[Rapid diagnosis of adenoviral conjunctivitis].

Adenoviral conjunctivitis is contagious and it should be diagnosed rapidly. Adenoviral ocular infection showed epidemic keratoconjunctivitis due to Ad8 and pharyngoconjunctival fever due to Ad3 but those clinical pictures are changing because of the appearance of new serotypes of adenovirus such as Ad4, 19 and 37 which are related with acute conjunctivitis. The diagnosis of viral infection is generally time consuming and complex. Enzyme-linked immunosorbent assay (ELISA) and immunofluorescent antibody (IFA) test, using monoclonal antibody to adenovirus hexon have been applied for rapid diagnosis. ELISA showed 75% sensitivity and 100% specificity in 59 cases with adenoviral conjunctivitis and was therefore shown to be more practical than IFA which showed only 67% sensitivity.

Detection of secretory IgM in tears during rhino-conjunctivitis.

BACKGROUND: With various methods, secretory immunoglobulin M (sIgM) was assessed in tears of patients with rhino-conjunctivitis. METHODS: Tears were analyzed by microimmunoelectrophoresis (MIE), size-exclusion high-pressure liquid chromatography (SE-HPLC), sodium dodecyl sulphate (SDS) electrophoresis and isoelectric focusing. RESULTS: Only very small traces of serum IgM could be found in tears of healthy volunteers. MIE showed that tear sIgM is different from serum IgM. The former migrates in the direction of the anode, while serum IgM migrates to the cathode. The SDS electropherogram of a number of patients showed an additional strong band of the sIgM mu-chain of approximately 68 kDa, after rhino-conjunctivitis. SE-HPLC showed two additional peaks for these patients, at 10.97 min and at 12.94 min, which were attributed to tear sIgM. The former peak consists of a complex of four sIgM molecules. One year later the chromatograms of the former rhino-conjunctivitis patients did not show any peak of sIgM in SE-HPLC. CONCLUSIONS: Serum IgM of a molecular weight (MW) 970 kDa appears only in normal tears at very low concentrations, as a result of transudation from the serum. In contrast, sIgM is synthesized during rhino-conjunctivitis in high concentrations in the human lacrimal glands and the conjunctiva. These increased levels of tear sIgM are due to stimuli for specific protein synthesis.

Mousepox conjunctivitis: the role of Fas/FasL-mediated apoptosis of epithelial cells in virus dissemination.

BALB/c mice infected with the Moscow strain of Ectromelia virus (ECTV-MOS) show a large number of apoptotic cells, and an influx of lymphoid cells in the epithelium and substantia propria of conjunctivae, respectively. The presence of ECTV-MOS antigens in the epithelium of conjunctivae significantly upregulates Fas in the epithelial layer and FasL in the suprabasal layer of conjunctiva. Inhibition of FasL with blocking antibodies in cultures of conjunctival cells isolated from ECTV-MOS-infected BALB/c mice showed that the Fas/FasL pathway is important in apoptosis of ECTV-MOS-infected cells. The results also showed that the presence of cytokines, in particular interferon (IFN)-gamma, upregulated expression of Fas. Interleukin (IL) 2, 4, 10 and IFN-gamma were produced at the peak of conjunctivitis (at day 15 of infection) with a predominance of IFN-gamma and a small, but significant, production of IL4 and IL10 compared with non-infected animals. These results suggest that not only is Fas/FasL expression in conjunctiva involved in elimination of migrating Fas+ cells but also plays an important role in the turnover of conjunctival epithelium and thus may be crucial for ECTV spreading to the surrounding environment.

The surgical treatment of Molluscum contagiosum in a pediatric AIDS patient.

Molluscum contagiosum (MC) has arisen as an opportunistic pathogen in human immunodeficiency virus (HIV) disease. Recent reports have focused on the presentation of MC in adults who are HIV positive. The authors describe MC in a pediatric patient with acquired immmune deficiency syndrome. This particular patient showed the same atypical lesion morphology, distribution, and chronic course that is seen in the adult HIV patient. As in the adult patient, treatment in the pediatric patient also remains challenging, with limited success of traditional treatment modalities. The authors were able to achieve long-term remission and an excellent cosmetic result through meticulous superficial surgical excision of the patient's MC lesions.

Comparison of direct rapid tests for the detection of adenovirus antigen in routine conjunctival specimens.

Two direct rapid tests to detect adenovirus antigen, enzyme immunoassay (EIA) and immunofiltration (IF) were compared with regard to sensitivity, specificity, ease of interpretation, and technical complexity against 75 adenovirus culture-positive and 35 adenovirus culture-negative conjunctival swab specimens. Enzyme immunoassay and IF were equally sensitive (P = 0.5), with sensitivities of 80.6 and 79.0%, respectively, when swab specimens were collected from patients within 7 days of the onset of clinical symptoms of adenovirus infection. After 7 days of clinical onset, IF (46.2%) was more sensitive (P less than 0.01) than EIA (0.0%), and was able to detect residual antigen in three culture-negative specimens. Enzyme immunoassay and IF were equally specific (100%). Both tests were easy to interpret, but IF was more technically complex and is not yet commercially packaged. Enzyme immunoassay could benefit either an office practice or a central laboratory, whereas IF is better suited for the latter.