Adult and neonatal inclusion conjunctivitis is caused primarily by the sexually transmitted disease (STD) strains of C. trachomatis. The historic link between these chlamydial strains and STDs, as well as the ocular diseases, was not made until diagnostics were developed for Neisseria gonorrhoeae (GC). Once it became clear that many cases of cervicitis, neonatal conjunctivitis, and nongonococcal urethritis could not be attributed to GC, the involvement of another infectious agent was suspected. In 1907, Halberstaedter and Prowazek⁴ identified typical chlamydial intracytoplasmic inclusions in conjunctival scrapings from an orangutan infected with a human trachoma strain. Similar inclusions were later seen in scrapings from the genital tracts of the parents of an infant with neonatal conjunctivitis.⁵ But it was not until 1959 that a strain responsible for chlamydial STDs was actually isolated. Because the natural history of chlamydial STDs is not well understood, including the propensity for latent or persistent infections to reactivate and be transmitted, the true incidence and prevalence of adult and neonatal inclusion conjunctivitis are not known. Adult inclusion conjunctivitis is usually a unilateral, self-limited disease of presumed low prevalence, although chlamydial genital strains are the most common cause of STDs in the developed and developing world today. This disease can also be caused by other species of Chlamydia such as Chlamydia psittaci and Chlamydia pneumoniae.^6–8 Neonatal conjunctivitis can be severe if untreated, and approximately 20% of infected neonates will develop pneumonitis within the first 6 months of life along with the long-term sequela of small airways disease in adulthood.

The last two decades have provided scientific advances in almost all areas of chlamydial research. These include clinical, diagnostic, epidemiologic, physiologic, pathologic, microbiologic, genetic, and immunologic characteristics of chlamydial diseases. Furthermore, the advent of a new, simplified trachoma grading scheme⁹ and a more specific strain typing technique referred to as ompA genotyping¹⁰ has enhanced the understanding of the clinical epidemiology, molecular epidemiology, transmission, risk factors, and prevalence of trachoma. However, the pathogenesis of trachomatous disease requires further study, and the best approach to designing an efficacious vaccine remains controversial.

With the exception of the rodent strains, C. trachomatis is currently known to infect only humans. The infections in humans include the conjunctiva and lower and upper genital tracts, including the rectum and lymphatics that drain the perineum. These infections are caused by the 19 currently recognized serologic variants (serovars) of the trachoma and LGV biovars. Serovars are defined by monoclonal and polyclonal antibodies that react to epitopes on the major outer membrane protein (MOMP) of C. trachomatis. Additional serovars presumably exist based on sequence data of the gene (ompA) that encodes for MOMP, but these strains have not been fully characterized to date.¹⁷ Serotyping has distinguished these serovars into different serogroups or classes: B class (serovars B, Ba, D, Da, E, L₂, and L_2a), intermediate class (serovars F and G), and C class (serovars A, C, H, I, Ia, J, Ja, K, Ka, L₁, and L₃).¹ Serovars A through K and Ba, Da, Ia, Ja, and Ka were previously referred to as trachoma-inclusion conjunctivitis (TRIC) strains.¹⁸ Trachoma is primarily caused by serovars A, B, Ba, and C, whereas adult and neonatal inclusion conjunctivitis are caused by serovars B or Ba, D through K, Da, Ia, Ja, Ka, L₁, L₂, L_2a, and L₃, which are the sexually transmitted strains of the organism. The LGV serovars tend to cause more severe disease and can invade regional lymphatics, whereas the non-LGV serovars are currently known to infect epithelial cells at ocular, respiratory, rectal, and genital mucosal surfaces.

Serotyping has been the most widely accepted technique for classifying C. trachomatis organisms. However, within the last decade, a new technique has been developed based on sequencing of ompA and is referred to as ompA genotyping.^1,19,20 (ompA was previously called omp1, but the nomenclature has changed to be consistent with that of other bacteria.) This latter technique has been and continues to be invaluable for evaluating the molecular epidemiology, disease pathogenesis, and transmission dynamics of chlamydiae for STD and trachoma populations.

New genus and species designations were proposed for chlamydiae in 1997 based on sequence analysis of small segments of rRNA.^21,22 However, this type of analysis provides relatively constant evolutionary relationships that do not necessarily translate into biologically important designations that help to distinguish one strain from another within a species or genus. This has become increasingly apparent from evidence of intragenic and intergenic recombination of the ompA gene for C. trachomatis, C. psittaci, and C. pneumoniae that affect persistence and tissue and host tropism for different stains.¹⁷ Although the rate of or time frame for mutations and recombination events is not known for ompA or other genes that encode for surface proteins on Chlamydia, a new taxonomy may not be appropriate because these events do occur. Furthermore, genomic analyses of other organisms have revealed biologic relationships that are not reflected in the phylogenies of rRNA-based sequences.²³ This continues to be an area of debate in the field of Chlamydia.

The developmental cycle is regulated by transcriptional events.²⁸ Cleavage of the disulfide bonds occurs within 6 to 8 hours of EB uptake and heralds the differentiation into the 800 to 1000 nm RB. The energy for RB replication, which occurs via binary fission starting at about 10 to 15 hours postinfection,²⁹ comes from its own stores of adenosine triphosphate (ATP) and activated ATPase, as well as essential metabolites and phosphate compounds from the host cell that are transported across the inclusion body membrane.³⁰ MOMP contains pore-like (porin) structures during the RB stage that are thought to facilitate this exchange.³¹ There appears to be a differential requirement for amino acids depending on the serovar: The LGV strains require methionine unlike the other STD strains, whereas the trachoma serovars require tryptophan. Leucine, phenylalanine, and valine, however, are necessary for growth for all known serovars and species of Chlamydia.³² As replication proceeds, the inclusion body expands and displaces the nucleus to the side of the cell. In addition, glycogen accumulates, which can be visualized by staining with iodine but is only present in C. trachomatis.

Within 36 to 72 hours, depending on the serovar, chromatin within the RB compacts down into an electron-dense nucleoid and, along with formation of membrane-bound disulfide bonds, produces an EB. The signal for these latter events is the reduction and oxidation of both intramolecular and intermolecular disulfide bonds.³¹ Anywhere from 100 to 1000 EBs can be produced per infected cell.³³ In many cases, the cell ruptures and dies releasing the infectious progeny, but the cell can also extrude the inclusion body by a process of exocytosis and, thereby, survive.

The EB is composed of a number of different proteins. MOMP comprises approximately 60% with a mass of 40 kDa³⁴ and is the most immunogenic of the surface proteins. The mass of this protein does differ somewhat according to the serovar. Both the lipopolysaccharide (LPS) and MOMP have genus-specific determinants, whereas MOMP also contains serovar-, serogroup-, and species-specific epitopes. There are four variable segments (VSs) interspersed by five constant regions on MOMP.³⁵ VS1, 2, and 4 are surface exposed and contain the serotyping epitopes of the organism, whereas VS3 has important T-cell determinants.^24,36 The genus-specific epitope of chlamydial LPS is a trisaccharide of 2-keto-3-deoxy-octanoic acid (KDO).³⁷ This approximately 7000 kDa glycolipid is similar to the LPS of enteric bacteria such as Salmonella typhimurium.³⁸ Both the LPS and MOMP are expressed throughout the developmental cell cycle, whereas the 12 and 60 kDa proteins discussed previously are expressed only at the time when the RB condenses back into an EB.²⁴

There are some important similarities and differences among the four species of Chlamydia. Although they differ in host and host cell preference, genomic composition, epitopes, drug susceptibility, and metabolism, they are similar in morphology and structure. However, the EB of C. pneumoniae can appear pear-shaped, instead of round, with a large periplasmic space. In addition, when more than one EB enters a cell, the inclusion bodies usually fuse into a single body but can remain distinct for C. pneumoniae, C. psittaci, and C. pecorum. The size of these bodies can be variable, especially for C. psittaci. Recently, nonfusing inclusions of C. trachomatis have been identified that have mutations in the inclusion membrane protein A (IncA) gene and lack IncA expression in the inclusion membrane.³⁹ Chlamydia contain a 7.5 kilobase (kb) plasmid that has been found only in C. trachomatis and C. psittaci strains. The sequence is relatively conserved within a species but varies considerably between the two species. The function of the plasmid is not known, although it is presumed to have a role in replication.⁴⁰ However, C. trachomatis strains that lack the plasmid can still be propagated in cell culture.⁴¹

Other genomic differences have been highlighted by the recent publications of the entire genome of C. trachomatis serovar D⁴²; C. pneumoniae strains CWL029, AR39, and J138^43,44; and C. psittaci strain GPIC (Read et al., unpublished data). The genome of C. pecorum has not yet been sequenced. Several open reading frames (ORFs) were found in each species that likely encode outer membrane proteins, including a polymorphic membrane protein (Pmp) gene (pmp) family including 9 genes for C. trachomatis, 21 for C. pneumoniae and 16 for C. psittaci. The pmp family is important because it is unique to the genus Chlamydia, comprises a surprising 10% of the genome, and interspecies amino acid sequence homology is less than 50% compared with 70% to 80% for other surface proteins. Although the function(s) of these proteins have not been determined, they are likely important in host immune response and pathogenesis. This is supported by some preliminary data that revealed differential immune responses to certain C. trachomatis Pmps in patients with chlamydial STDs.⁴⁵

All species of Chlamydia share the genus-specific epitope located on LPS. Serovar-, subspecies-, and species-specific epitopes on MOMP have been characterized for C. trachomatis and C. psittaci but do not appear to be immunogenic for C. pneumoniae. No data are available yet for C. pecorum. All species of Chlamydia are susceptible to tetracyclines, and all serovars of C. trachomatis are susceptible to sulfonamides. However, few strains of C. psittaci and C. pneumoniae strains respond to treatment with sulfonamides.

TRACHOMA

Trachoma is the leading infectious cause of preventable blindness in the world today and occurs as mesoendemic, endemic, or hyperendemic disease.¹ The distribution is primarily in tropical developing countries of the world including North and sub-Saharan Africa, the Middle East, and the Northern Indian subcontinent where the highest affliction rates are reported.² However, disease has also been reported in Southeast Asia and specific regions of Central and South America, Australia, and the South Pacific Islands.³ Of the more than 600 million people afflicted, approximately 150 million have visual deficits,³ and 12 million are predicted to be blind by the year 2020.⁴⁶ However, the precise prevalence of disease or disease severity is not known because surveys have not been conducted in many countries where trachoma is suspected nor have they been performed in many areas where trachoma is considered endemic or hyperendemic.

C. trachomatis is transmitted via eye to hand to eye contact where the primary reservoir of infection is the conjunctival mucosa. Other reservoirs include the nasopharynx and oropharynx and rectum, but their role in infection and reinfection of the ocular mucosa is not completely understood.⁴⁷ In one study, nasal secretions were not found to play a role in reinfection in a trachoma community in Tanzania,⁴⁸ but additional studies are required. The STD strains of C. trachomatis are also present in these communities but are not felt to contribute to trachoma. The incubation period is not known but is thought to range from 5 to 12 days based on nonhuman primate studies.⁴⁹ Children are considered the primary source of infection because they usually comprise more than 50% of the population and become easily infected from close contacts during frequent play and within small, crowded households. However, female caregivers can also serve as an important reservoir in which the infection is passed to them and then back to their children. Thus, children younger than 10 years are at greatest risk for infection and reinfection. Young women of childbearing age and other female caregivers are reported to have infection rates that range from 5% to 10%.^{47,48,50–52} These infections likely represent transmission within the household from and to children. Adult women develop more severe disease and sequelae than their male counterparts; repeat infection is considered an important factor in disease progression to trichiasis.

Patterns of trachomatous disease and disease severity are dependent on the degree of endemicity in villages or communities where C. trachomatis ocular infections occur. In hyperendemic areas, high prevalence rates of infection among children are found and reinfection is common. Indeed, most children by the age of 1 year have been infected at least once. Both sexes are equally at risk in childhood, but, because the rates of infection decline with age, adolescent and young adult females tend to have higher rates compared with their male counterparts. These communities have higher rates of conjunctival scarring, occurring as early as 4 years of age. Scarring is common in young adults and older age groups and progresses to trichiasis and entropion (in-turned eyelid) in many cases. These sequelae occur 10 to 40 years following the initial childhood disease⁵³; trichiasis rates range from 8% to 17.5% depending on the geographic locale.^54–56 Blindness occurs indirectly through corneal abrasion and bacterial superinfection, both of which may be facilitated by dry eyes, which is a common condition among older individuals. These lesions typically heal by scarring and fibrosis. Trachoma is the primary cause of blindness in hyperendemic areas.

In mesoendemic or endemic communities, trachoma takes two forms. In one, the prevalence is low but there are a number of households in which infection rates are high, which provides a reservoir for repeat infection that can lead to the sequelae mentioned previously. In the other form, infection occurs later, starting at school age, and blindness is rare. However, the community rate of infection may be moderate to high.^57,58 Where infection rates are declining, scarring in older individuals may still be present along with progression to trichiasis and blindness.

C. trachomatis cannot always be detected in all cases of active trachoma. In one report from Tanzania in which direct fluorescent antibody (DFA) and tissue culture tests were used, 43% of patients with follicular disease (TF) and 23% with intense inflammatory disease (TI) were negative for C. trachomatis.⁵⁰ Similar findings have also been reported for newer tests such as the ligase chain reaction (LCR)⁵⁹ and the polymerase chain reaction (PCR)⁶⁰ that have a higher sensitivity than DFA or culture. The most likely explanations for these findings are inadequate sampling of the conjunctiva, resolution of infection with residual inflammation, inflammation resulting from infections with other bacteria, or autoimmunity or some combination thereof.¹ In one study in The Gambia⁶¹ household members were tested for chlamydiae every 2 weeks. Organisms were detected 2 weeks before onset of inflammation and for an additional 2 to 6 weeks after disease development in children. Clinical disease persisted for 30 weeks in children and for only 2 weeks in adults. In other studies, 5% to 24% of villagers without active disease were found to be infected,^{47,48,50–52} including 24% of children in Tanzania⁵¹ who were PCR positive but had no evidence for trachoma. Thus, the time at which the sample is obtained can be critical for detection of the organism.

The degree of inflammatory disease varies considerably from one child to the next but can remain constant at different time points for a particular child despite treatment. Historically, environmental factors were considered to be responsible.⁶² However, a combination of factors is now thought to contribute to this phenomenon: environmental conditions, infectious load of C. trachomatis,⁶³ duration of infection,⁶⁴ host immune response,^1,65,66 and possibly host genetic susceptibility.^67,68 Severe inflammatory disease and the duration of inflammation in childhood is thought to be a risk factor for progression to scarring, and trichiasis and entropion later in life. Thus, age is also an important risk factor for trachomatous disease.⁶⁹ Additional interacting factors likely contribute. With age, there is continued exposure to the organism and risk of reinfection especially among females of childbearing age and female caregivers of all age groups. There can be changes in biologic susceptibility as in the development of dry eye syndrome that can occur from age or scarring. Also, host immune response to chlamydial antigens such as Hsp60 have been associated with trachomatous scarring, and this response may be potentiated with increasing age.⁶⁶

Other risk factors for trachoma are low socioeconomic status, poor facial hygiene, more than one child in a bedroom, many children per household, lack of water or lack of use of water, and proximity to cows.^50,70–72 Flies have historically been considered vectors for transmission in Africa. However, data from recent studies do not support the theory.⁷³ In one study, fluorescein was used to stain the secretions in the eyes of children.⁷⁴ Within 15 to 30 minutes, the legs and bodies of the flies were also stained with fluorescein. Eye-seeking flies such as Musca sorbens have been shown to land on the eyes of multiple children as was reported in a study in Africa,⁷⁵ but it is not clear how many infectious EBs can be carried on the flies, how long they are viable, and whether the inoculum is sufficient to cause infection. In The Gambia,⁷⁵ of 395 flies captured from the eyes of C. trachomatis-infected children, only two were positive by PCR and could not be confirmed. It is certainly possible that flies carry other bacteria from eye to eye, which might promote inflammatory disease and trachoma. Indeed, in many trachoma endemic countries, there are seasonal outbreaks of conjunctivitis resulting from multiple bacterial species including Haemophilus influenzae, Haemophilus aegyptius, Streptococcus pneumoniae, Neisseria meningitidis, N. gonorrhoeae, and Moraxella spp. These infections may actually precede periods of increased trachoma prevalence rates. In a study in Tunisia,⁵⁷ moderate to severe trachoma was found significantly more often among children with bacterial coinfections. Furthermore, pathogenic and nonpathogenic bacteria commonly colonize children who reside in trachoma areas. Coinfection of C. trachomatis with these bacteria may be one mechanism that is important for promoting severe inflammation, which results in conjunctival scarring and corneal vascularization years later.⁷⁶ In the study of trachoma pathogenesis, coinfection will be an important area to examine.

ADULT INCLUSION CONJUNCTIVITIS

Although chlamydial STDs account for more than 500 million cases worldwide with approximately 4 million annually in the United States,⁷⁷ the exact prevalence of adult inclusion conjunctivitis remains unknown, partly because infection is usually self-limited and does not always reach medical attention. The spread of chlamydial STD serovars is from hand to genital tract to eye contact or during sexual activity. However, there are reports of transmission occurring among family members from an infected neonate.¹⁸ Persons between the ages of 15 and 30 years are at highest risk for adult conjunctivitis.¹⁸ The incubation period is considered to be approximately 5 to 19 days, and disease may continue for months without treatment. There is usually complete resolution of disease following a 3-week course of systemic therapy with either tetracycline or erythromycin. Adult inclusion conjunctivitis is characterized by a serosanguineous to mucopurulent discharge with keratoconjunctivitis or a follicular inflammatory disease identical to trachoma. This clinical condition is not considered a precursor to the sequelae of trachoma. Cases that persist or in which reinfection occurs may lead to keratitis, corneal vascularization, and conjunctival scarring.

The LGV strains of C. trachomatis are responsible for a much more severe ocular disease referred to as Parinaud's oculoglandular syndrome.⁷⁸ This syndrome is comprised of an inflammatory conjunctival response with severe lymphadenopathy involving the preauricular, cervical, and submandibular nodes. The LGV serovars are uncommon in developed countries with few reports in the literature⁷⁹ but are very common in tropical and subtropical developing countries. Occasionally keratoconjunctivitis resulting from L₂ has been reported as a consequence of laboratory accidents.⁸⁰ The female cervix and rectum of homosexual men⁸¹ are important primary reservoirs of these infections.

NEONATAL INCLUSION CONJUNCTIVITIS

Approximately 5% of pregnant women have cervical infections with C. trachomatis.⁴⁹ If left untreated, there is a 50% chance that the infant will develop conjunctivitis. Neonatal inclusion conjunctivitis is about 10 times more common than conjunctivitis resulting from N. gonorrhoeae. The incubation period for chlamydial conjunctivitis is 1 to 3 weeks, unlike GC in which the incubation period is less than 7 days. Earlier infection can occur if there is evidence for rupture of membranes.⁸² If left untreated, the conjunctivitis can persist for 3 to 12 months. The outcome for late or no treatment is conjunctival scarring and corneal vascularization in 100% of these cases.⁸³ Infants born to an infected mother also carry a 20% to 30% chance of pneumonia. C. trachomatis is the leading cause of pneumonitis within the first 6 months of life. Rectal shedding of C. trachomatis does occur and is more common among infants with pneumonia. Onset of shedding does not usually occur before 6 to 12 weeks of age and can be as late as 12 months. This condition likely represents organisms that are swallowed after coughing as opposed to those that are acquired at birth. The short- and long-term importance of rectal shedding remains unclear.

TRACHOMA

Trachoma is a chronic follicular conjunctivitis persisting for 15 days or longer. It is characterized by follicles that cannot be distinguished from those caused by C. trachomatis STD strains, C. pneumoniae, or C. psittaci.⁷ Other entities also cause a follicular conjunctivitis including toxic follicular conjunctivitis resulting from cosmetics, drugs, or molluscum contagiosum; chronic follicular keratoconjunctivitis of Thygeson; folliculosis; Axenfeld's chronic follicular conjunctivitis; Vernal catarrh; Parinaud's oculoglandular syndrome; or other bacteria.⁵⁷ The latter bacteria are H. influenzae, H. aegyptius, Moraxella spp., N. meningitidis, N. gonorrhoeae, and S. pneumoniae.¹ The identification of C. trachomatis in suspected regions of trachoma endemicity provides good evidence that this is the etiologic agent despite the presence of other bacterial species or outbreaks of conjunctivitis.

The follicular stage of disease presents as a foreign body sensation and can resolve completely or progress to cicatricial conjunctivitis with superficial corneal vascularization. The lymphoid follicles are clear, yellowish or gray-white lesions of 0.2 to 2 mm in diameter in the subconjunctival epithelium primarily of the upper tarsal conjunctiva.⁵⁷ Papillary hypertrophy can occur, which is a result of inflammatory infiltrates. This finding may be the only sign of disease in children younger than age 2 years because follicle formation is not as common in this age group. Furthermore, infection in infants and young children may go undetected, thereby providing a source for transmission within the household and community.

The corneal involvement of the disease includes lymphoid follicle formation at the limbus.¹ This is a characteristic feature of trachoma and can lead to the development of Herbert's pits. Punctate epithelial erosions can occur anywhere on the cornea but are usually located at the superior corneal-scleral border. Other common corneal findings include anterior stroma infiltrates and shallow peripheral ulcers. These findings range from microscopic infiltrates to frank ulceration. Superficial corneal vessels, referred to as pannus, can also develop. Pannus can develop at any point along the limbal margin but is most pronounced at the superior limbal margin. Vision is not usually affected by pannus formation.

As the disease progresses, fine linear or stellate scars form horizontally across the upper tarsus. Children as young as 4 years can show signs of scarring. With age, the scars may expand into broad bands or, in severe cases, develop into synechial scars. As scarring progresses, especially toward the outer edge of the lid, the conjunctiva becomes distorted, resulting in trichiasis and entropion.¹ Corneal abrasions can occur from the inturned lashes and become superinfected with bacteria. In these cases, healing usually produces fibrosis and scarring. Thus, mild to severe corneal opacification can develop that can result in visual deficits or blindness. Loss of lid apposition and subsequent exposure keratitis or trauma may also develop with time. A late complication of scarring is lacrimal duct stenosis, which may lead to dry eye syndrome. This can also contribute to corneal ulceration, because there are few if any tears to flush out bacteria or to act as a barrier to infection. Because the severe sequelae of trachoma do not occur for 10 to 40 years following the initial infections, most of the visual deficits and blindness occur in the adult and older adult populations of trachoma endemic areas.

ADULT INCLUSION CONJUNCTIVITIS

Adults develop a follicular conjunctivitis that can be indistinguishable from that of trachoma. The follicles may be present on both the lower conjunctiva and upper tarsus. The onset is usually acute with preauricular lymphadenopathy on the involved side⁸⁴ and a serosanguineous to mucopurulent discharge. After 2 weeks of infection, corneal involvement is more prominent and includes keratitis, subepithelial opacities, and infiltrates that are marginal and/or central. Occasionally there is mild scarring and corneal vascularization referred to as micropannus,⁷⁴ but these are late findings, usually among cases that have not been treated. Otitis media is a common complication of chlamydial conjunctivitis.⁸⁵ Although there can be prompt resolution of the disease, conjunctivitis may persist for as long as 12 months if left untreated. In addition, failure to treat the genital tract disease can result in recurrence of conjunctivitis.

Conjunctivitis resulting from the LGV serovars of C. trachomatis cause a disease referred to as Parinaud's oculoglandular conjunctivitis. This clinical syndrome includes severe inflammation and lymphadenopathy of the preauricular, submandibular, and cervical nodes. These infections are uncommon in the United States. However, there are a number of reports dating back to the 1930s and 1940s.^78,86–89

NEONATAL INCLUSION CONJUNCTIVITIS

Conjunctivitis in the neonate is characterized by swelling of both lids, a purulent discharge, and hyperemia. Without treatment, neonates are at risk for conjunctival scarring, keratitis, and superficial vascularization of the cornea.⁹⁰ In addition, these infants are at risk for pneumonitis up to 6 months of age. The disease can persist for up to 12 months in untreated cases but does respond quickly to appropriate systemic treatment.

By adolescence or adulthood, the prevalence of active disease and the severity of the inflammatory response have diminished. For reasons that are not completely understood, some individuals develop conjunctival scarring that will progress to trichiasis and entropion years later, whereas others will not. This differential response to repeat infections is not explained by sociologic or environmental risk factors.⁹⁷ Usually only about 10% of adults older than age 25 years will have active disease, and the chlamydial isolation rates are approximately 1% for these cases.^94,98 Furthermore, many young adults harbor chlamydial antigens in their conjunctiva. Antigen presence is considered a risk factor for scarring disease.

Three theories are possible for the pathogenesis of trachomatous sequelae: (1) adults harbor persistent chlamydial organisms that usually cannot be isolated but provide a continuous or intermittent antigenic stimulus for deleterious host immune response (including the possibility that organisms may be associated with scarring but not causative); (2) after repeated infection, the organism is eliminated but chlamydial antigens (still present or now absent in tissue) trigger immune mediators that set up an initial inflammatory process that may or may not subside over the years and promote tissue destruction that results in scarring and trichiasis and entropion (includes the possibility that host genetic susceptibility plays a role in disease pathology); or (3) a combination of both.

Support for both theories comes from one study of American Indian school children in 1968 in whom active trachoma declined when the children attended a boarding school far from their endemic environment,⁹⁹ but chlamydial antigens still persisted in the ocular tissue of these children despite an absence of cultivable organisms.⁹⁹ Therefore, in addition to suggesting persistent infection, there may also be immune mediators that promote an immunopathogenic response that continues even when the individuals leave their endemic environment. In The Gambia, chlamydial antigens and secretory immunoglobulin A (sIgA) antibodies were found in 43% of adults who had conjunctival scarring but no evidence for active disease.⁹⁸ Elevated sIgA antibodies are not usually found at mucosal sites when there is no antigenic stimulation, as there would be from active infection. In the same study,⁹⁸ sIgA antibodies were present in 7% of adults who had no evidence for clinical disease and who resided in communities of low trachoma prevalence compared with 41% of adults living in high-prevalence communities. Also, tear IgG antibodies were significantly associated with antigen-positive adults compared with antigen-negative adults. These findings suggest that adults who are positive for chlamydial antigens have real yet unappreciated infection. These adults may harbor persistent organisms that intermittently replicate and produce antigen, thereby increasing the risk for fibrosis and scarring over subsequent years.

The ability of C. trachomatis to persist and evade the host immune response may be important in the pathogenesis of trachoma. Once infection is established, the organism can mutate at the gene and protein level in such a way that new immunogenic variant organisms (those derived from the infecting strain) arise that are able to evade the host immune response. In support of this is an in vitro study by Lampe and coworkers¹⁰⁰ in which neutralizing antibodies were shown to prevent infection for prototype strains, but closely related ompA gene variants escaped neutralization. There are a few studies that have described the occurrence of ompA mutants or variants that appear to have arisen from immune selection over time in trachoma endemic regions. The first such report was from Tunisia where 15 variants of 23 B and Ba serovars arose over a 3-year period in the same village area.¹⁰ Although the original serovars persisted in the community, a new immunologic serotype evolved during this period.¹⁹ Similar studies in The Gambia have also shown mutations in MOMP that have arisen over a 6-month interval,¹⁰¹ and a study in Tanzania revealed a high degree of ompA polymorphisms among trachoma serovars along with evidence for persistent infections.⁶³

Further evidence for in vivo persistence in trachoma populations comes from a study of conjunctival tissue obtained during surgery for trichiasis and for other types of corrective lid surgery.¹⁰² A significantly higher number of trichiasis patients were found to be positive for chlamydial DNA and RNA compared with the nontrichiasis patients, although all of the patients were from similar trachoma endemic regions of Vietnam. The tissue was examined for cell types by immunohistochemistry, and a predominance of macrophages was found in the trichiasis cases. Macrophages may function as cofactors both in inflammation and tissue injury. They induce tissue necrosis factor (TNF)-alpha but are also able to directly cause tissue injury through activation of complement (C3a) and hydrogen peroxide (H₂O₂) pathways and secretion of acid hydrolases.¹⁰³ Resident macrophages may phagocytose but not kill chlamydiae, thereby harboring the organism for a long time. Indeed, it has recently been shown that apoptosis is inhibited in C. trachomatis infected cells via inhibition of cytochrome c release from the mitochondria and caspase activation.¹⁰⁴ In another study, HeLa cells that were persistently infected with trachoma serovar A resisted apoptosis by mechanisms similar to those for acute infection.¹⁰⁵ High levels of Hsp60 with very low levels of MOMP and LPS were found in this study, which is consistent with in vitro studies of chlamydial persistence.¹⁰⁶ A recent study of chlamydial cervicitis revealed that approximately 5% of women had recurrent cervicitis, and 24% of these were with the same C. trachomatis serovar.¹⁰⁷ OmpA genotyping of recurrent samples from the same women showed identical genotypes or a few nucleotide changes that encoded for amino acid substitutions. Many intervening culture-negative samples were positive by LCR 4 weeks after treatment. Because 15 days is considered the maximum time that DNA can remain in tissue after elimination of viable organisms by antibiotic treatment,¹⁰⁸ these findings lend additional support to the growing body of evidence that chlamydial organisms can persist. Thus, chlamydiae may persist by using anti-apoptotic mechanisms, intermittently replicating, and producing antigen that elicits a deleterious host immune response.

As suggested previously, the host immune response to different antigens may also play a role in disease severity outcomes. Viable but not inactivated EBs, MOMP, or LPS can induce conjunctival inflammation within 24 hours. Another protein, the chlamydial Hsp60, which is a homologue of the 60 kDa GroEl Hsp of Escherichia coli, is also likely responsible for a direct inflammatory response.¹⁰⁹ Chlamydial Hsp60 is surface exposed and stimulates a vigorous humoral immune response in trachoma patients, as well as in patients with tubal infertility and ectopic pregnancy.^109–111 Furthermore, this protein has been associated with immunopathogenic responses in conjunctival^{105,109,112,113} and fallopian tube tissues.^111,114 Interestingly, in vitro studies of persistence induced by gamma interferon (IFN-gamma), penicillin,¹¹⁵ or amino acid deprivation¹¹⁶ have shown that viable organisms cannot be recovered during this persistent state. Furthermore, these infections upregulate Hsp60 production.¹⁰⁶ Because chlamydial Hsp60 has high homology with human Hsp60, it is possible that persistently infected cells trigger an autoimmune response to self-Hsp60.¹¹⁷ In a recent study of trachoma patients in Nepal,¹⁰⁵ specific tear anti-Hsp60 IgG immunoreactivity in C. trachomatis-infected patients was significantly associated with scarring disease, whereas high rates of anti-Hsp60 sIgA antibodies were not. However, it is not entirely clear whether Hsp60 directly causes disease, elicits a specific host immune response that is damaging, represents prolonged exposure to many different chlamydial antigens, or is a combination of these factors. In another study of in vitro persistence, cytokines and chemokines were released by persistent cells that may induce severe inflammation and fibrosis.¹¹⁸ In addition, a depressed CMI response may prevent the host from clearing an infection,¹¹⁹ which may lead to persistence, inflammation, and conjunctival scarring.¹ Thus, persistent organisms may chronically or intermittently produce immunogenic antigens or stimulate the release of cytokines/chemokines that precipitate the pathogenic responses seen in trachoma patients.

The role of genetic factors in determining severity of disease in trachoma populations is not well understood. In the mouse genital tract model of chlamydial STDs, the immune response to chlamydial Hsp60 was found to be genetically restricted.¹²⁰ This may in part explain the wide range in disease severity observed within trachoma endemic populations.¹ There are a few studies that suggest a genetic component to severity of the immune response in trachoma. An early study found that an important risk factor for scarring was evidence for maternal trichiasis.¹²¹ In another study, scarring was more common among individuals who had moderate to severe inflammation repeatedly in childhood.⁹⁴ It has also been reported that siblings tend to have the same degree of inflammatory response.⁶⁴ In the same study, persistently infected children had significantly more severe disease than children who were not persistently infected.⁶⁴ However, this may have been partially because the persistently infected children had significantly higher loads of chlamydial DNA. Nonetheless, these findings suggest that there is likely a genetic component to the host immune response in trachoma.

Specific genetic markers for disease susceptibility are ill defined and limited in geographic scope. HLA class I allele A*6802 has been associated with trachomatous scarring in one study in The Gambia.¹²² But the mechanism(s) of this association are unknown, although the finding would suggest a role for CD8 T cells in trachoma pathogenesis. Trachomatous scarring has also been associated with high titers of TNF-alpha in tears and with mutations in the promotor for TNF-alpha.⁶⁷ The potential mechanisms here include genetic determinants that control CD4 T_H1 versus T_H2 cytokine responses to chlamydiae and circulating antibody responses to chlamydial Hsp60. Other studies have shown elevated levels of IFN-gamma and interleukin (IL)-10 mRNA transcripts in severe trachomatous scarring,¹²³ suggesting a combined T_H1 and T_H2 immune response. In twin studies in The Gambia,¹²⁴ peripheral blood monocytes (PBMCs) were measured in response to chlamydial EBs and estimated at 39% (p = 0.07) for heritability. Additional studies of genetic markers in diverse trachoma populations worldwide will be needed to more fully estimate the role of host genetic determinants in chlamydial disease susceptibility.

The recently published sequence of the entire genome of C. trachomatis serovar D led to the identification of several ORFs predicted to encode outer membrane proteins, including a nine-member Pmp gene (pmp) family. Although it is not clear what role these membrane proteins may play in the pathogenesis of chlamydial ocular diseases, they are likely important because the pmp family is unique to the genus Chlamydia, comprises a surprising 10% of the genome, and interspecies amino acid sequence homology is less than 50% compared with 70% to 80% for other surface proteins. This will be an important and exciting area of research to define surface expression, antigenic determinants, and their potential role in host immune stimulation and pathogenesis.

The follicles can become necrotic with connective tissue formation surrounding the follicles that results in fibrosis.^1,129 As scarring occurs, posttrachomatous degeneration can develop. This is a condition in which the epithelium between papillae becomes trapped and forms cysts that are lined with epithelial cells. Follicles that form at the corneal-scleral junction do not cause ulceration or induce connective tissue formation, and, thus, there is no scar formation.¹ However, Herbert's pits can develop, which are visualized as depressed points usually at the superior limbal border.¹³⁰ Herbert's pits are the only pathognomonic feature of trachoma. Pannus formation can also occur at the limbal border and is characterized by a superficial vascularization of this tissue. It is usually localized to the superior limbus but can be appreciated anywhere along this border.¹ Trichiasis develops later on from gradual contraction of both the superficial and deep scars located near the lid margin.

Nonhuman primates have been experimentally inoculated with different strains of Chlamydia since the 1960s. Interestingly, the response to infection has varied depending on the primate species that was infected.^140–142 Chlamydial STD strains caused a severe follicular conjunctivitis in Old World monkeys (baboons, langurs, and Macaca spp.) who, in some cases, developed scarring and pannus. However, the latter was appreciated only in repeatedly infected Taiwan monkeys.¹⁴³ The trachoma serovars caused a milder form of the disease. The former infections resolved within 3 months at which time they were susceptible to reinfection. Trachoma serovar infections of New World monkeys (Aotus trivirgatus) resulted in acute conjunctivitis with disease resolution within 10 to 15 days and protective immunity to rechallenge.¹⁴¹

A model for chronic cicatrizing trachoma was produced in the cynomolgus monkey by weekly inoculations of either serovar A or E.⁹⁶ Both serovars produced similar results: a chronic follicular conjunctivitis with few cases of scarring but no pannus. Viable organisms could be cultured for up to 8 weeks. Peak systemic IgG and tear IgA antibodies were produced at the time of peak infection, although the tear IgA responses tended to develop more slowly and decrease more rapidly compared with the serum IgG responses. On conjunctival challenge, viable organisms were present only up to the time when IgA antibodies developed. A similar model in rhesus monkeys was not as successful.

The effect of initial infection with subsequent rechallenge, similar to what is seen among children in endemic regions, was also explored in the cynomolgus monkey model.⁹⁶ Following initial infection with serovar E, the monkeys developed a follicular conjunctivitis that persisted for 4 weeks with subsequent slow resolution during which time the organism could be cultured. At 15- to 30-weeks postinfection, the monkeys were rechallenged and found to develop a milder disease that resolved within 14 days. Thus, the animals were susceptible to reinfection, but the time to resolution was much shorter. The effect of immunosuppression on eye disease was also examined.¹⁴⁴ In these cases, monkeys were treated with cyclosporine and then infected with chlamydial serovars. The follicular conjunctivitis persisted and was associated with infection and lower levels of tear IgA levels compared with immunocompetent monkeys. Thus, local production of IgA appears to be important in protection and resolution of infection.

Oral immunization protocols with subsequent ocular challenge of chlamydial serovars were also investigated. Serovar B was used in some experiments, and no difference was appreciated for duration of disease or presence of viable organisms recovered from the conjunctiva.¹⁴⁵ Oral immunization with LGV followed by serovar B inoculation of the conjunctiva revealed a milder conjunctivitis but prolonged shedding of viable organisms. In contrast, use of ultraviolet-inactivated L₂ for oral immunization resulted in prolonged disease and shedding. Thus, nonviable organisms were not able to stimulate an appropriate immune response. Immunization by different routes (oral, intramuscular, and rectal) resulted in early resolution of disease and shedding.

The impact of CMI response has also been studied in the cynomolgus monkey model. During peak infection, the conjunctiva of these animals were sectioned and stained for T-cell phenotypes.¹⁴⁶ Both CD8+ and CD4 cells were present, but the former predominated, which suggested that a DTH response was likely involved.

In the early murine model of chlamydial ocular infection, limited duration of infection and lymphoproliferative responses were observed, although the mice did develop systemic IgG antibodies.¹³² More recently, three inbred strains of mice were infected with serovar C of C. trachomatis applied to the conjunctiva.¹³³ Two strains of mice, BALB/c and C3H/HeN, were susceptible to infection. The organism could be recovered for up to 21-days postinfection, and histopathologic analyses revealed subepithelial mononuclear infiltration and loss of goblet cells within 1 week of infection. Furthermore, lymphoproliferative responses to chlamydial EBs were present for infected but not for uninfected mice. This model has been used recently to test an orally and systemically administered monoclonal anti-idiotypic antibody (anti-Id) to chlamydial exoglycolipid antigen (GLXA) as a vaccine to stimulate systemic immunity.¹⁴⁷ Protection from ocular challenge with a C. trachomatis human serovar occurred, producing systemic anti-chlamydial neutralizing antibodies and anti-GLXA antibodies. However, there were no data regarding mucosal IgA responses, which are likely important in disease prevention and resolution. The mouse model of trachoma, however, is potentially exciting, because it can take advantage of the vast array of resources available for studies in mice, as well as the body of knowledge that has been accumulating over the last decade from the mouse model of chlamydial genital tract infections.

CLINICAL CONSIDERATIONS

A presumptive diagnosis of trachoma can be made based on clinical features, especially in an area where trachoma is considered to be present. The following signs are important indicators for trachoma, and at least two must be present per person examined¹⁴⁸: follicles in the upper tarsal conjunctiva, limbal follicles or their sequelae, Herbert's pits (which are pathognomonic for trachoma), typical conjunctival scarring, and vascular pannus. These signs along with conjunctival detection of C. trachomatis confirm the presence of endemic or hyperendemic trachoma in the respective area.

The World Health Organization (WHO) developed an extensive grading scale for trachoma, the MacCallan classification system,¹⁴⁸ which has subsequently been modified for use in field applications.⁹ This modification has assisted in the understanding of the epidemiology of clinical disease, risk factors for disease, and disease prevalence.⁶¹ The simplified trachoma grading scale includes

  TF: trachoma follicles: 5 or more follicles in the central upper tarsal conjunctiva
  TI: intense trachoma: diffuse infiltration of the upper tarsal conjunctiva that obscures the deep tarsal vessels over 50% or more of the tarsal surface
  TS: trachomatous scarring of the conjunctiva
  TT: trichiasis (inturned eyelashes) that touches the eyeball
  CO: corneal opacity that impairs vision
  Active trachoma: presence of TF or TI
  Intense trachoma: presence of TI
  Cicatricial trachoma: presence of TS
  Healed trachoma: TS without TF or TI

Although this grading scale retains some features of the older system, including intensity of inflammation (TF, TI), trichiasis (TT), and sequelae (CO), it does not go into detail, for example, on the degree of conjunctival scarring that is essential for predicting disease complications.⁶¹

CYTOLOGY

Conjunctival smears or swabs are required for making a diagnosis. The tests available to detect C. trachomatis include Giemsa stain, Lugol's iodine, or DFA for smears and for swabs, culture, enzyme-linked immunosorbent assay (ELISA), DNA probes, or commercial PCR or LCR. The conjunctiva is swabbed with a Dacron or cotton swab, and the smear is made by rolling the swab over a clean glass slide. Alternatively, the swab can be placed in a special transport media for the respective diagnostic test. Conjunctival smears should first be examined for the quality of the specimen. Epithelial cells that are clearly separated and the presence of PMNs, lymphocytes, plasma cells, or Leber cells (giant macrophages that contain phagocytosed material) can denote an adequate sampling of the conjunctiva. The degree of inflammation and bacterial superinfection can also be appreciated from these smears. Thus, although not diagnostic for chlamydiae, these findings are suggestive of trachoma.¹⁴⁹ Giemsa stain is inexpensive, and the test is easy to perform, which makes it attractive for developing countries where trachoma is endemic or hyperendemic. However, the sensitivity is only about 60% and, thus, should not be used in areas of low endemicity.¹ With this stain, the inclusion body is visualized as a basophilic, stippled inclusion in contrast to the dark blue to purple color of the cell. However, other entities can also stain similarly: These include goblet cells, bacteria, keratin, nuclear extrusions, and eosinophilic granules.¹⁴⁹ Lugol's iodine stains the glycogen-containing inclusion of C. trachomatis. It imparts a dark yellow-brown color to the inclusion but is infrequently used, because it is insensitive. The DFA test detects the EBs instead of the inclusion body. Commercially available fluorescent [fluorescein isothiocyanate (FITC)] conjugated monoclonal antibodies against the MOMP, which is species-specific for C. trachomatis, or the LPS, which is genus-specific for Chlamydia, are used in this test.¹⁵⁰ The EBs are stained an apple green color and are visualized as extracellular round dots. Technical expertise is critical to differentiate debris from true EBs. The sensitivity for this test is approximately 80% to 90%.

TISSUE CULTURE

Although the intracellular inclusions of trachoma were first identified by Halberstaedter and von Prowazek⁴ in 1907, the actual organisms were finally cultured in 1957 by using chick embryos.¹⁵¹ Today, tissue culture has supplanted the use of eggs, which has made isolation of chlamydiae more widely available, although it is still only performed in specialized reference laboratories. Tissue culture remains the gold standard for C. trachomatis identification but is not 100% sensitive,¹⁵² probably because of the difficulty in maintaining a cold chain (4°C for no longer than 24 hours and then -70°C) from the field site to a specialized reference lab where the culture will actually be performed. Also, because some viability is lost on freezing and some of the trachoma serovars are more difficult to propagate, the enthusiasm for this test is decreased. Furthermore, culture requires technical expertise, can take 3 to 6 days for results, and is very expensive. Many different cell lines are now available for culture, including HeLa and McCoy cells. Additional passages in tissue culture can increase the positive rate but have other drawbacks, including a delay in the reporting time of the results. Fluorescein-conjugated antibodies are used to detect the inclusions in cell culture, which are visualized as intracytoplasmic ovoid, round, or irregularly shaped inclusions. This stain imparts a fluorescing, apple green color to the inclusion body that stands out against the dark red cells that have been counterstained with Evans blue. Peroxidase-conjugated monoclonal antibodies are also available for the detection of chlamydial inclusion bodies but are less frequently used.

ANTIGEN DETECTION

ELISA or enzyme immunoassay (EIA) commercial assays are available to detect chlamydiae, but the sensitivity is only 70% to 85%.¹ However, these assays can be cost effective compared with other commercially available tests such as the DNA detection assays¹⁵³(see later). These tests detect the EB via polyclonal or monoclonal antibodies directed against the genus-specific chlamydial LPS. The antibodies are conjugated with an enzyme that reacts with a substrate to produce a change in color that can be detected by a specific wavelength in a spectrophotometer. One advantage is that a 96-well format can be used to process multiple samples at one time. Less technical expertise is required than for the above-mentioned tests. Another advantage is that the kits contain a confirmatory test. The chlamydial LPS is blocked by an antibody, and the sample is rerun; if a positive sample is now negative, then it is considered a true positive. However, if a positive sample is still positive, then it is a false positive, because the assay is probably detecting LPS from other bacterial species.

DNA DETECTION

The commercial LCR (Abbott) and PCR (Roche Diagnostics) tests are the most recent assays to be developed for detecting Chlamydia. Primers that are specific for the organism anneal to the complementary strand of DNA after denaturation. This target DNA is usually the plasmid, which is only present in C. trachomatis and C. psittaci species. There have been some recent reports of C. trachomatis strains that do not contain the plasmid.¹⁵⁴ Exactly how many strains are missing the plasmid is not known; the numbers are likely to be small and so would not significantly affect the sensitivity of these tests, which are in the range of 92% to 99%. LCR amplifies a signal that occurs when the primers hybridize with the plasmid DNA. In PCR, the actual DNA is amplified after hybridization. Both tests can be used in a 96-well format in which 92 to 94 samples can be assayed at one time. Both products are detected by spectrophotometers that are set at specific wavelengths for the particular assay. An advantage to the commercial PCR test is that an internal control plate can be run in parallel with the chlamydial detection plate to identify which samples have inhibitors. Those samples that contain inhibitors can then be run by in-house PCR assays that employ a DNA purification protocol that removes the inhibitors. However, the internal control plates do add to the cost of the assay, and not all labs have the capabilities to perform the in-house purification assays or PCR. The commercial tests have been used in very few trachoma studies to date^59,155,156 but appear to have similar sensitivities and specificities as in STD populations.

Chlamydial DNA can also be detected by commercially available hybridization probes. These also hybridize with complementary plasmid or ompA DNA. The sample is usually a swab of the conjunctiva that has been applied to a special filter paper immediately after the sample has been obtained from the patient. Occasionally, DNA is extracted from a swab that has been placed in a special collection media and then is applied to a filter. In both cases, the filter is what is probed. The advantage of this technique is that the filter paper that contains the samples can be stored at room temperature under field conditions and transported back to the lab at a convenient time, without the necessity of a cold chain. The sensitivity of the probes is 70% to 90%.

SEROLOGY

There are two serologic tests for Chlamydia: the microimmunofluorescent (MIF) test¹⁵⁷ and the complement fixation (CF) test. However, neither is specific for the organism because patient sera can crossreact with different serovars and species and may represent current or previous sexually transmitted infection as opposed to conjunctival infection. The highest antibodies detected in the assay, however, are usually found against the initial infecting serovar, even on subsequent infection. This concept is referred to as original antigenic sin.¹⁵⁸ Furthermore, ocular chlamydial infections tend to be chronic and endemic. Thus, these assays cannot be used to diagnose active infection, although occasionally MIF has been used for epidemiologic studies. They are also only available in reference laboratories.

The CF test is the older of the two and detects group-reactive antigen on C. psittaci and C. trachomatis. This test can be used for diagnosing ocular infections resulting from LGV or C. psittaci. The MIF test employs EBs representing C. trachomatis serovars and usually one or two strains of C. pneumoniae and C. psittaci. Sera, tears, and other bodily fluids can be used in this assay. The fluids are serially diluted and reacted against the EBs that have been applied and fixed to a slide in groups of dots. A FITC conjugated antihuman IgM, IgG, IgA or secretory IgA antibody is used as the secondary antibody to detect antigen-antibody binding. The slides are screened under fluorescent microscopy for fluorescing EBs that represent the respective serovar or species. Serum IgM and IgG antibodies appear around 2- to 3-weeks postinfection and persist for 4 to 8 weeks, although the IgG antibodies persist for much longer. Occasionally, IgM titers can rise again with reinfection or relapse of infection. Approximately 80% of children in trachoma endemic areas¹⁵⁹ and 90% of adults with inclusion conjunctivitis¹⁶⁰ will have detectable MIF antibodies. About half of the population in trachoma endemic areas will have both serum and tear antibodies, and the titers are directly proportional to the severity of disease and to the presence of chlamydial organisms in the conjunctiva. In one study in Tunisia, 80% of children with severe disease, 31% with moderate disease, and 17% with mild disease had tear antibodies to chlamydiae.¹⁵⁹ Although the highest titers are usually against the infecting serovar in ocular infections, the MIF test can be used only for a diagnosis of active infection in neonates not in older children or adults. Neonates acquire IgG antibodies from their mothers, and when they develop conjunctivitis, these titers usually do not change during the course of the ocular infection. Most infected neonates do develop a small rise in serum IgM antibodies, usually less than 1:32, which persists for a few weeks.¹⁵⁷

TRACHOMA

Trachoma was initially treated using copper sulfate solutions or crystals. The introduction of sulfonamides allowed for their use in the treatment of American Indian populations, which resulted in a marked decline in trachoma prevalence rates.⁸³ For many tribes, trachoma was eradicated by 1942. Most developed countries have experienced complete elimination of trachoma because of a combination of factors including improved personal hygiene, improved sanitation, and access to clean water. Because of the lack of these socioeconomic indicators in developing countries, where trachoma is endemic, the WHO initiated control programs that used topical treatment with tetracyclines and erythromycins. Topical treatment, which was applied as continuous or intermittent regimens over a period of weeks or months, resulted in disease reduction, but these effects were short lived. Longer term and more consistent success rates were achieved with systemic treatment using sulfonamides or tetracyclines.¹⁶¹ However, a major problem with these regimens was compliance with taking a drug multiple times for many days. Also, children and pregnant women could not be treated with tetracycline drugs because it is contraindicated. A newer azalide antibiotic, azithromycin, has recently been introduced and used to treat trachoma cases in Africa. One to six doses of oral azithromycin, administered either as a community-wide treatment program or to selected persons, resulted in reduction of active trachoma and infection rates for approximately 12 months.^{59,155,162,163} However, no long-term follow-up was done to determine the time interval at which infection rates began to rise or returned to baseline. It may be possible that multiple treatments of azithromycin over a few years will prove to lower the rates of infection so that the burden of disease is decreased and blindness no longer occurs. But more comprehensive studies will need to be done to determine if this is likely. Furthermore, studies will need to address whether resistance to azithromycin develops as a consequence of mass treatment programs. To date, resistance to tetracycline and erythromycin has been reported but only for sporadic cases of sexually transmitted strains of C. trachomatis.¹⁶⁴

Because azithromycin and mass treatment programs are expensive, along with the unlikely possibility that the socioeconomic status of trachoma-endemic countries will change anytime soon, one is still faced with the challenge of controlling trachoma to prevent blinding disease. The number of persons who are blind from trachoma is estimated to be 12 million by the year 2020.⁴⁶ The WHO initiated the Global Elimination of Trachoma 2020 program (GET 2020), which employs the SAFE strategy.^2,165 The SAFE program encompasses four different arms of treatment and prevention of disease: surgery for TT; antibiotics for active disease; face washing and good personal hygiene; and environmental improvements, consisting of improved sanitation (waste disposal and better water delivery systems). A number of trachoma-endemic countries in Africa and Asia have instituted the SAFE program, but results of these programs will not be available for a few years.

ADULT INCLUSION CONJUNCTIVITIS

The best form of treatment for adult inclusion conjunctivitis is to prevent chlamydial STDs. Unfortunately, most chlamydial STDs are asymptomatic for males (approximately 40%) and females (approximately 70%) and usually go undetected because routine diagnostic screening for C. trachomatis is not performed.¹ Thus, it is important to recognize adult inclusion conjunctivitis that is caused by C. trachomatis and treat both the ocular and genital tract disease. Because chlamydial STDs cannot be resolved by topical ocular antibiotics, systemic therapy is recommended. Most cases infected with non-LGV serovars will respond to 2 to 3 weeks of oral tetracycline, doxycycline, or erythromycin.¹⁶⁶ For LGV cases, 6 weeks is required for complete eradication.¹⁶⁷ The best treatment regimen for inclusion conjunctivitis caused by C. psittaci and C. pneumoniae is unknown, although 6 weeks of oral antibiotics has been successful in some cases.⁷

NEONATAL INCLUSION CONJUNCTIVITIS

Treatment of pregnant women is effective in preventing 90% of infants from acquiring conjunctival infection,¹⁶⁸ although retreatment may be necessary in high-risk populations.¹⁶⁹ Topical treatment of neonatal inclusion conjunctivitis does not eradicate nasopharyngeal carriage, which can result in pneumonia or recurrent ocular infection. Thus, systemic treatment is recommended. Oral erythromycin at 50 mg per kg of body weight in four divided doses is required for 10 to 14 days. Approximately 80% of cases will respond to this regimen, but additional treatment may be necessary for some cases.¹⁷⁰ In addition, mothers should be treated for genital tract infection to prevent recurrence of chlamydial conjunctivitis in the neonate.

1. Dean D. Trachoma. In Connor DH, Chandler FW, Manz HJ et al. (eds): Pathology of Infectious Diseases. Stamford, Conn.: Appleton and Lange. 1997:498–507

2. Thylefors B, Negrel AD, Pararajasegaram R et al. Global data on blindness. Bull World Health Org 1995;73:115

3. World Health Organization. Report of WHO: Future approaches to trachoma control. Report of a Global Scientific Meeting, Geneva, 1996

4. Halberstaedter L, von Prowazek S. Zur Atiologie des trachoma. Dtsch med Wochenschr 1907;33:1285

5. Lindner K. Gnoblennorrhoe, einschlussblennorrhoe und trachoma. Graefes Arch Clin Exp Ophthalmol 1911;78:380

6. Lietman T, Brooks D, Moncada J et al. Chronic follicular conjunctivitis associated with Chlamydia psittaci or Chlamydia pneumoniae. Clin Infect Dis 1998;26:1335

7. Dean D, Shama A, Schachter J et al. Molecular identification of an avian strain of Chlamydia psittaci causing severe keratoconjunctivitis in a bird fancier. Clin Infect Dis 1995;20:1179

8. Ostler HB, Schachter J, Dawson CR. Acute follicular conjunctivitis of epizootic origin. Feline pneumonitis. Arch Ophthalmol 1969;82:587

9. Thylefors B, Dawson CR, Jones BR et al. A simple system for the assessment of trachoma and its complications. Bull World Health Org 1987;65:477

10. Dean D, Schachter J, Dawson CR et al. Comparison of the major outer membrane protein variant sequence regions of B/Ba isolates: A molecular epidemiologic approach to Chlamydia trachomatis infections. J Infect Dis 1992;166:383

11. Weisburg WG, Hatch TP, Woese CR. Eubacterial origin of chlamydiae. J Bacteriol 1986;167:570–574.

12. Moulder JW, Hatch JP, Kuo J et al. Bergey's Manual of Systematic Bacteriology. Vol 1. Baltimore: Williams and Wilkins 1984:729–739

13. Grayston JT, Kuo CC, Campbell LA et al. Chlamydia pneumoniae sp. nova for chlamydia sp. strain TWAR. Int J Sys Bacteriol 1989;39:88

14. Pudjiatmoko, Fukushi H, Ochiai Y et al. Phylogenetic analysis of the genus Chlamydia based on 16S rRNA gene sequences. Int J Syst Bacteriol 1997;47:425

15. Nigg C. An unidentified virus which produces pneumonia and systemic infection in mice. Science 1942;95:49

16. Stills HF, Fox JG, Paster BJ et al. A “new” Chlamydia sp. strain SFPD isolated from transmissible proliferative ileitis in hamsters. Microbiol Ecol Health Dis 1991;4:S99

17. Millman KL, Tavare S, Dean D. Recombination in the ompA gene but not the omcB gene of Chlamydia contributes to serovar-specific differences in tissue tropism, immune surveillance, and persistence of the organism. J Bacteriol 2001;183:5997

18. Jones BR. Ocular syndromes of TRIC virus infection and their possible genital significance. Br J Vener Dis 1964;40:13

19. Dean D. Molecular characterization of new Chlamydia trachomatis serological variants from a trachoma endemic region of Africa. Chlamydial Infections. Bologna, Italy: Societa Editrice Esculapios. 1994:259–262

20. Dean D, Millman K. Molecular and mutation trends analyses of omp1 alleles for serovar E of Chlamydia trachomatis.Implications for the immunopathogenesis of disease. J Clin Invest 1997;99:475

21. Everett KD, Andersen AA. The ribosomal intergenic spacer and domain I of the 23S rRNA gene are phylogenetic markers for Chlamydia spp. Int J Syst Bacteriol 1997;47:461

22. Everett KD, Bush RM, Andersen AA. Emended description of the order Chlamydiales, proposal of Parachlamydiaceae fam. nov. and Simkaniaceae fam. nov., each containing one monotypic genus, revised taxonomy of the family Chlamydiaceae, including a new genus and five new species, and standards for the identification of organisms. Int J Syst Bacteriol 1999;49(Pt 2):415

23. Pennisi E. Genome data shake tree of life. Science 1998;280:672

24. Morrison RP, Manning DS, Caldwell HD. Immunology of Chlamydia trachomatis infections: Immunoprotective and immunopathogenic responses. Sex Transm Dis 1992;8:57

25. Zhang JP, Stephens RS. Mechanism of C. trachomatis attachment to eukaryotic host cells. Cell 1992;69:861

26. Vretou E, Goswami PC, Bose SK. Adherence of multiple serovars of Chlamydia trachomatis to a common receptor on HeLa and McCoy cells is mediated by thermolabile protein(s). J Gen Microbiol 1989;135:3229

27. Wyrick PB, Choong J, Davis CH et al. Entry of genital Chlamydia trachomatis into polarized human epithelial cells. Infect Immun 1989;57:2378

28. Jones RB. Chlamydial Diseases. In Mandell GL, Bennett JE, Dolin R (eds): Principles and Practice of Infectious Diseases. New York: Churchill Livingstone. 1995:1676–1693

29. Alexander JJ. Separation of protein synthesis in meningopneumonitis agent from that in L cells by differential susceptibility to cycloheximide. J Bacteriol 1968;95:327

30. Tipples G, McClarty G. The obligate intracellular bacterium Chlamydia trachomatis is auxotrophic for three of the four ribonucleoside triphosphates. Mol Microbiol 1993;8:1105

31. Bavoil P, Ohlin A, Schachter J. Role of disulfide bonding in outer membrane structure and permeability in Chlamydia trachomatis. Infect Immun 1984;44:479

32. Pearce JH, Allan I. Differential amino acid requirements of chlamydiae: Regulation of growth in relationship with clinical syndrome. In Mardh PA, Homes KK, Oriel JD (eds): Chlamydia Infections. Amsterdam: Elsevier Biomedical Press. 1982:29–32

33. Moulder JW. Interaction of chlamydiae and host cells in vitro. Microbiol Rev 1991;55:143

34. Caldwell HD, Kromhout J, Schachter J. Purification and partial characterization of the major outer membrane protein of Chlamydia trachomatis. Infect Immun 1981;31:1161

35. Stephens RS, Sanchez-Pescador R, Wagar EA et al. Diversity of Chlamydia trachomatis major outer membrane protein genes. J Bacteriol 1987;169:3879

36. Su H, Morrison RP, Watkins NG et al. Identification and characterization of T helper cell epitopes of the major outer membrane protein of Chlamydia trachomatis. J Exp Med 1990;172:203

37. Brade H, Brade L, Nano FE. Chemical and serological investigations on the genus-specific lipopolysaccharide epitope of Chlamydia. Proc Natl Acad Sci U S A 1987;84:2508

38. Nurminen M, Leinonen M, Saikku P et al. The genus-specific antigen of Chlamydia: Resemblance to the lipopolysaccharide of enteric bacteria. Science 1983;220:1279

39. Suchland RJ, Rockey DD, Bannantine JP et al. Isolates of Chlamydia trachomatis that occupy nonfusogenic inclusions lack IncA, a protein localized to the inclusion membrane. Infect Immun 2000;68:360

40. Hatt C, Ward ME, Clarke IN. Analysis of the entire nucleotide sequence of the cryptic plasmid of Chlamydia trachomatis serovar L1. Evidence for involvement in DNA replication. Nucleic Acids Res 1988;16:4053

41. Peterson EM, Markoff BA, Schachter J et al. The 7.5-kb plasmid present in Chlamydia trachomatis is not essential for the growth of this microorganism, Plasmid 1990;23:144

42. Stephens RS, Kalman S, Lammel C et al. Genome sequence of an obligate intracellular pathogen of humans: Chlamydia trachomatis. Science 19998;282:754

43. Grimwood J, Stephens RS. Computational analysis of the polymorphic membrane protein superfamily of Chlamydia trachomatis and Chlamydia pneumoniae. Microb Comp Genomics 1999;4:187

44. Shirai M, Hirakawa H, Kimoto M et al. Comparison of whole genome sequences of Chlamydia pneumoniae J138 from Japan and CWL029 from USA. Nucleic Acids Res 2000;28:2311

45. Hsia RC, Ahmed I, Batteiger B et al. Differential immune response to polymorphic membrane proteins in STD patients. Fourth Meeting of the European Society for Chlamydia Research. Helsinki: Societa Editrice Esculapio. 2000:219

46. Schachter J, Dawson CR. The epidemiology of trachoma predicts more blindness in the future. Scand J Infect Dis 1990;69(Suppl):55

47. Malaty R, Zaki S, Said ME et al. Extraocular infections in children in areas with endemic trachoma. J Infect Dis 1981;143:853

48. West S, Munoz B, Bobo L et al. Nonocular Chlamydia infection and risk of ocular reinfection after mass treatment in a trachoma hyperendemic area. Invest Ophthalmol Vis Sci 1993;34:3194

49. Stamm WE, Holmes KK. Chlamydia trachomatis infections of the adult. In Holmes KK, Mardh PA, Sparling PF et al. (eds): Sexually Transmitted Diseases. New York: McGraw-Hill. 1990:181–193

50. Taylor HR, Rapoza PA, West S et al. The epidemiology of infection in trachoma. Invest Ophthalmol Vis Sci 1989;30:1823

51. Bobo L, Munoz B, Viscidi R et al. Diagnosis of Chlamydia trachomatis eye infection in Tanzania by polymerase chain reaction/enzyme immunoassay. Lancet 1991;338:847

52. Dean D, Palmer L, Pant CR et al. Use of a Chlamydia trachomatis DNA probe for detection of ocular chlamydiae. J Clin Microbiol 1989;27:1062

53. Kupka K, Nizetic B, Reinhards J. Sampling studies on the epidemiology and control of trachoma in southern Morocco. Bull World Health Org 1968;39:547

54. Munoz B, Bobo L, Mkocha H et al. Incidence of trichiasis in a cohort of women with and without scarring. Int J Epidemiol 1999;28:1167

55. Rabiu MM, Abiose A. Magnitude of trachoma and barriers to uptake of lid surgery in a rural community of northern Nigeria. Ophthalmic Epidemiol 2001;8:181

56. Khandekar R, Mohammed AJ, Courtright P. Recurrence of trichiasis: A long-term follow-up study in the Sultanate of Oman. Ophthalmic Epidemiol 2001;8:155

57. Schachter J, Dawson CR. Trachoma. Human Chlamydial Infections. Littleton, Colo: PSG Publishing. 1978;63–96

58. Assaad FA, Sundaresan T, Maxwell-Lyons F. The household pattern of trachoma in Taiwan. Bull World Health Org 1971;44:605

59. Schachter J, West SK, Mabey D et al. Azithromycin in control of trachoma. Lancet 1999;354:630

60. Bailey RL, Hayes L, Pickett M et al. Molecular epidemiology of trachoma in a Gambian village. Br J Ophthalmol 1994;78:813

61. Dawson CR. Trachoma and other chlamydial eye diseases. In Orfila J, Byrne GI, Chernesky MA (eds): Chlamydial Infections. Bologna, Italy: Societa Editrice Esculapio. 1994:277–287

62. Dawson CR, Daghfous T, Messadi M et al. Severe endemic trachoma in Tunisia. Br J Ophthalmol 1976;60:245

63. Hsieh YH, Bobo LD, Quinn TC et al. Determinants of trachoma endemicity using Chlamydia trachomatis ompA DNA sequencing. Microbes Infect 2001;3:447

64. Bobo LD, Novak N, Munoz B et al. Severe disease in children with trachoma is associated with persistent Chlamydia trachomatis infection. J Infect Dis 1997;176:1524

65. Ghaem-Maghami S, Bailey RL, Mabey DC et al. Characterization of B-cell responses to Chlamydia trachomatis antigens in humans with trachoma. Infect Immun 1997;65:4958

66. Hessel T, Dhital SP, Plank R et al. Immune response to chlamydial 60-kilodalton heat shock protein in tears from Nepali trachoma patients. Infect Immun 2001;69:4996

67. Conway DJ, Holland MJ, Bailey RL et al. Scarring trachoma is associated with polymorphism in the tumor necrosis factor alpha (TNF-alpha) gene promoter and with elevated TNF-alpha levels in tear fluid. Infect Immun 1997;65:1003

68. White AG, Bogh J, Leheny W et al. HLA antigens in Omanis with blinding trachoma: Markers for disease susceptibility and resistance. Br J Ophthalmol 199781:431

69. Bailey R, Osmond C, Mabey DC et al. Analysis of the household distribution of trachoma in a Gambian village using a Monte Carlo simulation procedure. Int J Epidemiol 1989;18:944

70. West SK, Rapoza P, Munoz B et al. Epidemiology of ocular chlamydial infection in a trachoma-hyperendemic area. J Infect Dis 1991;163:752

71. West SK, Congdon N, Katala S et al. Facial cleanliness and risk of trachoma in families. Arch Ophthalmol 1991;109:855

72. Brechner RJ, West S, Lynch M. Trachoma and flies. Individual vs environmental risk factors. Arch Ophthalmol 1992;110:687

73. Mabey D, Fraser-Hurt N. Trachoma. BMJ 2001;323:218

74. Jones BR. The prevention of blindness from trachoma. Trans Ophthalmol Soc U K 1975;95:16

75. Emerson PM, Bailey RL, Mahdi OS et al. Transmission ecology of the fly Musca sorbens, a putative vector of trachoma. Trans R Soc Trop Med Hyg 2000;94:28

76. Dawson CR, Whitcher JP, Lyon C et al. Response to treatment in ocular chlamydial infections (trachoma and inclusion conjunctivitis): Analogies with nongonococcal urethritis. In Hobson D, Holmes KK (eds): Nongonococcal Urethritis. Washington DC: Am. Soc. Microbiol. 1977:135–139

77. Jones RB, Batteiger BE. Chlamydia Trachomatis. In Mandell GL, Bennett GE, Dolin R (eds): Douglas and Bennett's Principles and Practice of Infectious Diseases, Ed 5. Philadelphia: Churchill Livingstone. 2000:1986-1989

78. Curth W, Curth HO, Sanders M. Chronic conjunctivitis due to the virus of venereal lymphogranuloma. JAMA 1941;115:445

79. Buus DR, Pflugfelder SC, Schachter J et al. Lymphogranuloma venereum conjunctivitis with a marginal corneal perforation. Ophthalmology 1988;95:799

80. Rake G, Shaffer MS, Thygeson P. Relationship of the agents of trachoma and lymphogranuloma psittacosis group. Proc Soc Exp Biol Med 1942;49:545

81. Quinn TC, Goodell SE, Mkrtichian E et al. Chlamydia trachomatis proctitis. N Engl J Med 1981;305:195

82. Schachter J, Grossman M, Holt J et al. Prospective study of chlamydial infection in neonates. Lancet 1979;2:377

83. Forster RK, Dawson CR, Schachter J. Late follow-up of patients with neonatal inclusion conjunctivitis. Am J Ophthalmol 1970;69:467

84. Jones BR, Hussani K, Dunlop E. Infection of the eye and genital tract by TRIC agent. III. Ocular syndromes associated with infection of the genital tract by TRIC agent. Revue du Trachome 1965;42:27

85. Gow JA, Ostler HB, Schachter J. Inclusive conjunctivitis with hearing loss [Letter]. JAMA 1974;229:519

86. Bollock J, Basch G, Desvignes P. Infection Conjonctivale d'adenopathie du au virus du la maladie du Nicholas-Favre. Bull Soc Optalmol Paris 1936;409

87. Levaditi C. Conjonctivite avec adenopathie du au virus du la maladie du Nicholas-Favre: Lymphogranulomatose a localisation oculaire. Bull Soc Franc Dermatol Syphillol 1936;43:1238

88. Applemans M. Conjonctivite infectious de Parinaud. Ophthalmologica 1939;96:321

89. Oliphant JW, Pound WF, Perrint L. Sulfadiazine in lymphogranuloma venereum ophthalmitis. JAMA 1942;118:973

90. Mordhorst CH, Dawson C. Sequelae of neonatal inclusion conjunctivitis and associated disease in parents. Am J Ophthalmol 1971;71:861

91. Grayston JT, Wang SP, Yeh LJ et al. Importance of reinfection in the pathogenesis of trachoma. Rev Infect Dis 19857:717

92. Grayston JT, Woolridge RL, Wang SP. Trachoma vaccine studies in Taiwan. Ann N Y Acad Sci 1962;98:352

93. Wang SP, Grayston JT, Alexander ER. Trachoma vaccine studies in monkeys. Am J Ophthalmol 1967;63:1615

94. Mabey DC, Bailey RL, Ward ME et al. A longitudinal study of trachoma in a Gambian village: Implications concerning the pathogenesis of chlamydial infection. Epidemiol Infect 1992;108:343

95. Monnickendam MA, Darougar S, Treharne JD et al. Development of chronic conjunctivitis with scarring and pannus, resembling trachoma, in guinea-pigs. Br J Ophthalmol 1980;64:284

96. Taylor HR, Johnson SL, Prendergast RA et al. An animal model of trachoma II. The importance of repeated reinfection. Invest Ophthalmol Vis Sci 1982;23:507

97. West SK, Munoz B, Lynch M et al. Risk factors for constant, severe trachoma among preschool children in Kongwa, Tanzania. Am J Epidemiol 1996;143:73

98. Ward M, Bailey R, Lesley A et al. Persisting inapparent chlamydial infection in a trachoma endemic community in The Gambia. Scand J Infect Dis 1990;69(Suppl):137

99. Hanna L, Dawson CR, Briones O et al. Latency in human infections with TRIC agents. J Immunol 1968;101:43

100. Lampe MF, Kuehl LM, Wong KG et al. Chlamydia trachomatis major outer membrane protein variants escape neutralization by polyclonal human immune sera. In Orfila J, Byrne GI, Chernesky MA et al (eds): Chlamydial Infections. Bologna, Italy: Societa Editrice Esculapio. 1994:91–94

101. Hayes LJ, Pecharatana S, Bailey RL et al. Extent and kinetics of genetic change in the omp1 gene of Chlamydia trachomatis in two villages with endemic trachoma. J Infect Dis 1995;172:268

102. Phan S, Pham R, Dean D. Characterization of the local immune response to Chlamydia trachomatis among trachoma patients with conjunctival scarring. New York: International Symposium on Immunopathogenesis of Chlamydial Infections. 1996:198(abst)

103. Roitt IM. Adversarial strategies during infection. In Roitt IM (ed): Essential Immunology. Oxford: Blackwell Scientific Publications. 1994:243–271

104. Fan T, Lu H, Hu H et al. Inhibition of apoptosis in chlamydia-infected cells: Blockade of mitochondrial cytochrome c release and caspase activation. J Exp Med 1998;187:487

105. Dean D, Powers VC. Persistent Chlamydia trachomatis infections resist apoptotic stimuli. Infect Immun 2001;69:2442

106. Beatty WL, Byrne GI, Morrison RP. Morphologic and antigenic characterization of interferon gamma-mediated persistent Chlamydia trachomatis infection in vitro. Proc Natl Acad Sci U S A 1993;90:3998

107. Dean D, Suchland R, Stamm W. Evidence for long-term cervical persistence of Chlamydia trachomatis by omp1 genotyping. J Infect Dis 2000;182:909

108. Gaydos CA, Crotchfelt KA, Howell MR et al. Molecular amplification assays to detect chlamydial infections in urine specimens from high school female students and to monitor the persistence of chlamydial DNA after therapy. J Infect Dis 1998;177:417

109. Morrison RP, Lyng K, Caldwell HD. Chlamydial disease pathogenesis. Ocular hypersensitivity elicited by a genus-specific 57-kD protein. J Exp Med 1989;169:663

110. Bierly J, Dawson CR, Jones M et al. Serological evaluation for 60,000 molecular weight heat shock protein (HSP 60) in patients with trachoma. Invest Ophthalmol Vis Sci 1992;33:848

111. Wagar EA, Schachter J, Bavoil P et al. Differential human serologic response to two 60,000 molecular weight Chlamydia trachomatis antigens. J Infect Dis 1990;162:922

112. Morrison RP, Belland RJ, Lyng K et al. Chlamydial disease pathogenesis. The 57-kD chlamydial hypersensitivity antigen is a stress response protein. J Exp Med 1989;170:1271

113. Taylor HR, Maclean IW, Brunham RC et al. Chlamydial heat shock proteins and trachoma. Infect Immun 1990;58:3061

114. Toye B, Laferriere C, Claman P et al. Association between antibody to the chlamydial heat-shock protein and tubal infertility. J Infect Dis 1993;168:1236

115. Beatty WL, Morrison RP, Byrne GI. Persistent chlamydiae: From cell culture to a paradigm for chlamydial pathogenesis. Microbiol Rev 1994;58:686

116. Coles AM, Reynolds DJ, Harper A et al. Low-nutrient induction of abnormal chlamydial development: A novel component of chlamydial pathogenesis? FEMS Microbiol Lett 1993;106:193

117. Yi Y, Yang X, Brunham RC. Autoimmunity to heat shock protein 60 and antigen-specific production of interleukin-10. Infect Immun 1997;65:1669

118. Rasmussen SJ, Eckmann L, Quayle AJ et al. Secretion of proinflammatory cytokines by epithelial cells in response to Chlamydia infection suggests a central role for epithelial cells in chlamydial pathogenesis. J Clin Invest 1997;99:77

119. Holland MJ, Bailey RL, Hayes LJ et al. Conjunctival scarring in trachoma is associated with depressed cell-mediated immune responses to chlamydial antigens. J Infect Dis 1993;168:1528

120. Zhong G, Brunham RC. Antibody responses to the chlamydial heat shock proteins hsp60 and hsp70 are H-2 linked. Infect Immun 1992;60:3143

121. Turner VM, West SK, Munoz B et al. Risk factors for trichiasis in women in Kongwa, Tanzania: A case-control study. Int J Epidemiol 1993;22:341

122. Conway DJ, Holland MJ, Campbell AE et al. HLA class I and II polymorphisms and trachomatous scarring in a Chlamydia trachomatis-endemic population. J Infect Dis 1996;174:643

123. Holland MJ, Bailey RL, Conway DJ et al. T helper type-1 (Th1)/Th2 profiles of peripheral blood mononuclear cells (PBMC); responses to antigens of Chlamydia trachomatis in subjects with severe trachomatous scarring. Clin Exp Immunol 1996;105:429

124. Bailey R, Fowler A, Peeling R et al. Lymphoproliferative responses to C. trachomatis EBS in a Gambian twin population; estimating the role of host genetic factors. Chlamydial Infections. Proceedings of the Ninth International Symposium on Human Chlamydial Infection. Napa, Calif: International Chlamydial Symposium. 1998:474–477

125. Cosgrove PA, Patton DL, Kuo CC et al. Experimentally induced ocular chlamydial infection in infant pig-tailed macaques. Invest Ophthalmol Vis Sci 1989;30:995

126. el-Asrar AM, Van den Oord JJ, Geboes K et al. Immunopathology of trachomatous conjunctivitis. Br J Ophthalmol 1989;73:276

127. Patton DL, Taylor HR. The histopathology of experimental trachoma: Ultrastructural changes in the conjunctival epithelium. J Infect Dis 1986;153:870

128. Badir G, Wilson RP, Maxwell-Lyons F. The histopathology of trachoma. Bull Egypt Ophthalmol Soc 1953;46:129

129. Duke-Elder S. Diseases of the outer eye. In Duke-Elder S (ed): System of Ophthalmology. London: Henry Kingston. 1965;VIII:578

130. Dawson CR, Juster R, Marx R et al. Limbal disease in trachoma and other ocular chlamydial infections: Risk factors for corneal vascularisation. Eye 1989;3:204

131. Murray ES. Guinea pig inclusion conjunctivitis virus. Isolation and identification as a member of the psittacosis-lymphogranuloma-trachoma group of organisms. J Infect Dis 1964;114:1

132. Barsoum IS, Hardin LK, Colley DG. Immune responses of mice after conjunctival exposure to Chlamydia trachomatis serovar A. Med Microbiol Immunol Berl 1988;177:349

133. Whittum-Hudson JA, O'Brien TP, Prendergast RA. Murine model of ocular infection by a human biovar of Chlamydia trachomatis. Invest Ophthalmol Vis Sci 1995;36:1976

134. Malaty R, Dawson CR, Wong I et al. Serum and tear antibodies to Chlamydia after reinfection with guinea pig inclusion conjunctivitis agent. Invest Ophthalmol Vis Sci 1981;21:833

135. Watson RR, MacDonald AB, Murray ES et al. Immunity to chlamydial infections of the eye. 3. Presence and duration of delayed hypersensitivity to guinea pig inclusion conjunctivitis. J Immunol 1973;111:618

136. Modabber F, Bear SE, Cerny J. The effect of cyclophosphamide on the recovery from a local chlamydial infection. Guinea-pig inclusion conjunctivitis (GPIC). Immunology 1976;30:929

137. Watson RR, Mull JD, MacDonald AB et al. Immunity to chlamydial infections of the eye. II. Studies of passively transferred serum antibody in resistance to infection with guinea pig inclusion conjunctivitis. Infect Immun 1973;7:597

138. Orenstein NS, Mull JD, Thompson SE III. Immunity to chlamydial infections of the eye. V. Passive transfer of antitrachoma antibodies to owl monkeys. Infect Immun 1973;7:600

139. Nichols RL, Murray ES, Nisson PE. Use of enteric vaccines in protection against chlamydial infections of the genital tract and the eye of guinea pigs. J Infect Dis 1978;138:742

140. Dawson CR, Jawetz E, Thygeson P et al. Trachoma virus isolated in the United States. IV. Immunogenicity for monkeys. Porc Soc Exp Biol Med 1961;106:898

141. Nichols RL, Oertley RE, Fraser EC et al. Immunity to chlamydial infections of the eye. VI. Homologous neutralization of trachoma infectivity for the owl monkey conjuctivae by eye secretions from humans with trachoma. J Infect Dis 1973;127:429

142. Wang SP, Grayston JT. Pannus with experimental trachoma and inclusion conjunctivitis agent infection of Taiwan monkeys. Am J Ophthalmol 1967;63(Suppl):1133

143. Wang SP. Clinical evaluation of monkey eye infection with TRIC agent. A numerical scoring system of disease severity. Am J Ophthalmol 1967;63(Suppl):1321

144. Young E, Schachter J, Prendergast RA et al. The effect of cyclosporine in chlamydial eye infection. Curr Eye Res 1987;6:683

145. Taylor HR, Young E, MacDonald AB et al. Oral immunization against chlamydial eye infection. Invest Ophthalmol Vis Sci 1987;28:249

146. Whittum-Hudson JA, Taylor HR, Farazdaghi M et al. Immunohistochemical study of the local inflammatory response to chlamydial ocular infection. Invest Ophthalmol Vis Sci 1986;27:64

147. Whittum-Hudson JA, An LL, Saltzman WM et al. Oral immunization with an anti-idiotypic antibody to the exoglycolipid antigen protects against experimental Chlamydia trachomatis infection. Nat Med 1996;2:1116

148. Dawson CR, Jones BR, Tarizzo ML. Guide to Trachoma Control. Geneva: World Health Organization. 1981:7–56

149. Yoneda C, Dawson CR, Daghfous T et al. Cytology as a guide to the presence of chlamydial inclusions in Giemsa-stained conjunctival smears in severe endemic trachoma. Br J Ophthalmol 1975;59:116

150. Tam MR, Stamm WE, Handsfield HH et al. Culture-independent diagnosis of Chlamydia trachomatis using monoclonal antibodies. N Engl J Med 1984;310:1146

151. T'ang FF, Wang HI, Chang YT. Studies on the etiology of trachoma with special reference to isolation of the virus in chick embryo. Chin Med J 1957;75:429

152. Dean D, Pant CR, O'Hanley P. Improved sensitivity of a modified polymerase chain reaction amplified DNA probe in comparison with serial tissue culture passage for detection of Chlamydia trachomatis in conjunctival specimens from Nepal. Diagn Microbiol Infect Dis 1989;12:133

153. Dean D, Ferrero D, McCarthy M. Comparison of performance and cost-effectiveness of direct fluorescent-antibody, ligase chain reaction, and PCR assays for verification of chlamydial enzyme immunoassay results for populations with a low to moderate prevalence of Chlamydia trachomatis infection. J Clin Microbiol 1998;36:94

154. Farencena A, Comanducci M, Donati M et al. Characterization of a new isolate of Chlamydia trachomatis which lacks the common plasmid and has properties of biovar trachoma. Infect Immun 1997;65:2965

155. Holm SO, Jha HC, Bhatta RC et al. Comparison of two azithromycin distribution strategies for controlling trachoma in Nepal. Bull World Health Organ 2001;79:194

156. Baral K, Osaki S, Shreshta B et al. Reliability of clinical diagnosis in identifying infectious trachoma in a low-prevalence area of Nepal. Bull World Health Organ 1999;77:461

157. Wang SP, Grayston JT, Alexander ER et al. Simplified microimmunofluorescence test with trachoma-lymphogranuloma venereum (Chlamydia trachomatis) antigens for use as a screening test for antibody. J Clin Microbiol 1975;1:250

158. Wang SP, Grayston JT. Microimmunofluorescence antibody responses in Chlamydia trachomatis infection: A review. In Mardh PA, Holmes KK, Oriel JD (eds): Chlamydial Infections. Amsterdam: Elsevier Biomedical Press. 1982:301

159. Treharne JD, Dwyer RS, Darougar S et al. Antichlamydial antibody in tears and sera, and serotypes of Chlamydia trachomatis isolated from schoolchildren in Southern Tunisia. Br J Ophthalmol 1978;62:509

160. Hanna L, Jawetz E, Briones OC et al. Antibodies to TRIC agents in tears and serum of naturally infected humans. J Infect Dis 1973;127:95

161. Dawson CR, Hanna L. A resume of experience in controlled trials with topical and systemic tetracycline and systemic sulfonamide. Ophthalmol Dig 1971;33:30

162. Dawson CR, Schachter J, Sallam S et al. A comparison of oral azithromycin with topical oxytetracycline/polymyxin for the treatment of trachoma in children. Clin Infect Dis 1997;24:363

163. Bailey RL, Arullendran P, Whittle HC et al. Randomised controlled trial of single-dose azithromycin in treatment of trachoma. Lancet 1993;342:453

164. Jones RB, Van der Pol B, Martin DH et al. Partial characterization of Chlamydia trachomatis isolates resistant to multiple antibiotics. J Infect Dis 1990;162:1309–1315

165. World Health Organization. Report of the First Meeting of the WHO Alliance for the Global Elimination of Trachoma. Geneva: World Health Organization. 1997

166. Viswalingam ND, Darougar S, Yearsley P. Oral doxycycline in the treatment of adult chlamydial ophthalmia. Br J Ophthalmol 1986;70:301

167. National guideline for the management of lymphogranuloma venereum. Clinical Effectiveness Group (Association of Genitourinary Medicine and the Medical Society for the Study of Venereal Diseases). Sex Transm Infect 1999;75(Suppl 1):S40

168. Crombleholme WR, Schachter J, Grossman M et al. Amoxicillin therapy for Chlamydia trachomatis in pregnancy. Obstet Gynecol 1990;75:752

169. Oh MK, Cloud GA, Baker SL et al. Chlamydial infection and sexual behavior in young pregnant teenagers. Sex Transm Dis 1993;20:45

170. Pereira LH, Embil JA, Haase DA et al. Cytomegalovirus infection among women attending a sexually transmitted disease clinic: Association with clinical symptoms and other sexually transmitted diseases. Am J Epidemiol 1990;131:683

171. Batteiger BE, Rank RG, Bavoil PM et al. Partial protection against genital reinfection by immunization of guinea-pigs with isolated outer-membrane proteins of the chlamydial agent of guinea-pig inclusion conjunctivitis. J Gen Microbiol 1993;139:2965

172. Pal S, Theodor I, Peterson EM et al. Immunization with an acellular vaccine consisting of the outer membrane complex of Chlamydia trachomatis induces protection against a genital challenge. Infect Immun 1997;65:3361

173. Zhang D, Yang X, Berry J et al. DNA vaccination with the major outer-membrane protein gene induces acquired immunity to Chlamydia trachomatis (mouse pneumonitis) infection. J Infect Dis 1997;176:1035

174. Zhang D, Yang X, Shen C et al. The immune responses following intramuscular DNA immunization with the MOMP gene of Chlamydia trachomatis. Chlamydial Infections. Proceedings of the Ninth International Symposium on Human Chlamydial Infection. San Francisco: International Chlamydia Symposium. 1998:442–445

175. Su H, Messer R, Whitmire W et al. Vaccination against chlamydial genital tract infection after immunization with dendritic cells pulsed ex vivo with nonviable Chlamydiae. J Exp Med 1998;188:809

176. Brunham RC. Vaccine design for the prevention of Chlamydia trachomatis infection. In Orfila J, Byrne GI, Chernesky MA et al (eds): Chlamydial Infections. Proceedings of the Eighth International Symposium on Human Chlamydial Infections. Bologna, Italy: Societa Editrice Esculapio. 1994:73–82