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1 Review

2 Title of manuscript: Infectious keratoconjunctivitis in small ruminants with special

3 emphasis to Mycoplasma conjunctivae

4 Author: Dr. Md. Ahaduzzaman

5 Affiliation:

6 1. Md. Ahaduzzaman, Assistant Professor, Department of Medicine & Surgery, Chittagong

7 Veterinary & Animal Sciences University, Khulshi-4202, Chittagong, Bangladesh.

8 Email: zaman.cvasu@gmail.com

9 Abbreviation note list

10 DGGE: Denaturing gradient gel electrophoresis

11 ELISA: Enzyme-linked immunosorbent assay

12 I/M: Intra-muscular

13 IBR: Infectious bovine rhinotracheitis

14 IKC: Infectious keratoconjunctivitis

15 Kg: Kilogram

16 LppS: Lipoprotein S

17 LppT: Lipoprotein T

18 Mg: Milligram

19 PCR: Polymerase chain reaction

20 PPR: Peste des Petits Ruminants

21 S/C: Sub-cutaneous

22 TVH: Teaching veterinary hospital

23 UV: Ultra violet

24
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25 Abstract

26 Infectious keratoconjunctivitis (IKC) is a widespread and important ocular

27 manifestation of small ruminants, characterized by variable degrees of corneal opacity and

28 blindness. Though it has a culminating prognosis, there is a gap between the knowledge of

29 infection and therapeutics for IKC. Studies have mainly focused on wild rather than

30 domestic habitats. Therapeutic feasibility may not be feasible in the open range, but could

31 have values in domestic areas where small ruminants are considered as the “poor man’s

32 cow” in agribusiness. This article will address the general aspects of IKC in small

33 ruminants to overview potential values in the treatment and control.

34 Keywords

35 Pink eye, mycoplasmosis, ocular treatment, animal welfare

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37

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44 1. Introduction

45 Infectious keratoconjunctivitis (IKC), commonly known as pink eye disease, is an

46 infectious and highly contagious ocular infection, frequently reported in domestic sheep

47 and goats, and in wild ruminants (1). Mycoplasma is considered to be the primary

48 pathogen causing ocular inflammation without involvement of secondary microorganisms.

49 In a previous survey on keratoconjunctivitis-affected herds in Norway to find out the

50 causative agent of IKC in sheep, Mycoplasma conjunctivae was isolated from 37% of

51 animals with clinical signs and from 7% of those without clinical signs (2). The main route

52 of transmission of IKC between and within flocks appears to be the introduction to a

53 disease free flock of one or more animals such as (e.g. a ram) with either a subclinical

54 infection or unapparent carrier status (3). As with other infectious disease, susceptibility to

55 IKC varies, and several risk factors are thought to increase susceptibility. These factors

56 include exposure to high intense sunlight, sand and dust storms, fighting wound, poor

57 feeding, housing and health care management (4).

58 Inflammation of the conjunctiva results in lacrimation, blinking or blepharospasm.

59 The disease may progress and involve the cornea, sometimes causing extensive keratitis.

60 In such cases, vision is seriously impaired. The infection is sometimes confused with

61 presence of foreign bodies, parasites or trauma within the eyes. However, response to

62 treatment is thought to be a good indicator for diagnosis of IKC (3). Furthermore,

63 laboratory confirmation is mainly based on biomolecular methods (PCR, DGGE) which

64 are rapid and accurate, and overcome the problems related to isolation of the causative

65 organism. Since animals require special treatment and some nursing to reduce the period

66 of blindness, farmers may see IKC as a nuisance (5). Prophylaxis requires a quarantine

67 period for all newly introduced animals, and appears to be the reliable way to prevent the
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68 spread of the disease (6).

69 There is a considerable knowledge gap in relation to M. conjunctivae infections in

70 small ruminants, particularly in the case of sheep and goats. Therefore, this review gives

71 an overview of IKC and emphasizes the importance of future research into the disease, in

72 areas where small ruminants are an important part of the economy.

73 2. Aetiology and host range

74 The term IKC is used to describe a clinical condition, but the causative agent has

75 been a topic of debate for a long time. Several etiological agents have been proposed,

76 including Moraxella ovis (7), Chlamydia psittaci (8), Corynebacterium pyogenes, M.

77 conjunctivae, Staphylococcus aureus, Neisseria ovis (9), Streptococcus spp., Escherichia

78 coli, Pseudomonas spp. (10), Listeria monocytogenes (11), Rickettsia (12), IBR virus and

79 fungus (13). However, in domestic sheep, pathogenicity has been demonstrated for C.

80 psittaci and M. conjunctivae, with these organisms having been isolated from the eyes of

81 sheep infected with IKC (14). Pathogenicity of M. conjunctivae has also been documented

82 in the domestic goat (15), Alpine ibex (16) and European mouflon; also, this agent has

83 been implicated as the etiological agent of IKC in Alpine chamois (17). M. conjunctivae is

84 now generally agreed to be the principal single cause of IKC in domestic and wild

85 Caprinae (18). Based on genomic structure, M. conjunctivae appears to belong to the M.

86 neurolyticum cluster of the hominis group, which is closely related to M. ovipneumoniae

87 and M. bovoculi (19). M. conjunctivae were first isolated from sheep’s eyes by Surman in

88 Australia in 1968, and were later described and named by Barile et al. (20). M.

89 conjunctivae have been isolated from domestic sheep and goats in Australia, North

90 America, Europe, Saudi Arabia and the South Africa (14, 21-23). Stress, ocular foreign
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91 bodies and arthropods may act as risk factors and aggravate the condition (24).

92 3. Epidemiology

93 Most of the epidemiological studies related to M. conjunctivae infection have been

94 carried out in mountain habitats. Prevalence may present year round, but peak infections

95 are common in late spring (May–June), early fall (September–October) (23) and summer

96 (25, 26). Transmission of infection maybe either direct or indirect. Direct transmission is

97 mainly through nasal and ocular droplets. Indirect transmission has been documented as

98 occurring through the house fly (Musca domestica) (17), face fly (Musca autumnalis) or

99 stable fly (Stomoxys calcitrans) (27). Interspecies transmission may occur (28), and

100 domestic sheep seem to play a key role as a reservoir host (29). Ear mites (Raillietia

101 caprae) in the external ear canal may be linked to infection (30) but their role in the

102 transfer of mycoplasma between animals is unknown (31).

103 In domestic sheep and goats, some outbreaks of IKC have been described. The

104 prevalence of conjunctivitis varies according to the underlying cause, but has been

105 reported to be between 10% and 76%, and may also be influenced by the animal’s

106 immune status, age and sex, and by the season of the year (32).The immunity to

107 mycoplasma associated with keratoconjunctivitis is poor, and recovered animals may act

108 as a carrier for that particular herd (33). Although the morbidity rate may be high depends

109 on causative agent, mortality rates are usually negligible. Ovine keratoconjunctivitis

110 typically causes more severe clinical signs in adult sheep than in lambs (34) and infection

111 may be due to a previous Peste des Petits Ruminants (PPR) outbreak (35). Some breeds

112 appear to be less susceptible, but the extent to which genetic selection and breeding will

113 decrease the incidence of infection is unknown (36). Animals with pigmented skin around
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114 the eye generally seem more resistant to infection than those that are non-pigmented. M.

115 conjunctivae may be viable in the conjunctival sac for several months; also, it may be

116 harboured in the nares of animals, providing opportunities for re-infections within a flock

117 (18, 37). Elevation enhance the disease course in both ibex and chamois; this could be due

118 to altitude-associated environmental conditions such as increased UV radiation, cold and

119 dryness (38).

120 4. Pathogenesis

121 It is expected that M. conjunctivae have efficient adhesion functions in order to

122 attach in the presence of lacrimal secretion. Adhesion is thought to play a central role in

123 the pathogenicity, both directly as a basic condition of colonization (39-42) and indirectly

124 by adherence coupled to cytopathic functions. In relation to such functions, adhering

125 mycoplasmas may induce the release of peroxide and oxygen radical species to the

126 targeted cell, resulting in oxidative damage to the host cell (43, 44). Extracellular matrix

127 proteins such as lipoprotein T (LppT), lipoprotein S (LppS) and glycosaminoglycans are

128 important receptors for adhesion of pathogens (45). Following adhesion, mycoplasmas are

129 known to induce biological events such as cell differentiation, malignant transformation,

130 immune recognition and blood coagulation (Table 1) (46, 47). Although the actual

131 molecular mechanism is unclear, it has reported that three proteins toxins – hemolysin A

132 (hlyA), hemolysin B (hlyB) and hemolysin C (hlyC)– are important virulence factors

133 scattered within the M. conjunctivae genome (48).

134 5. Clinical signs

135 The severity of clinical signs and possible clearance or persistence of infection

136 varies (49). Incubation may be as short as 1–3 days or as long as 3 weeks, and may occur
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137 unilaterally or bilaterally (50). Animals frequently exhibit blepahrospasm; that is,

138 hyperaemia of conjunctiva with purulent discharge (51). Gradually, the cornea may

139 become opaque and this may spread centrifugally over the entire corneal surface (52)

140 (Figure 1). In a small number of cases, perforation of the corneal ulcers may be seen; this

141 results in iris prolapse and permanent blindness. Secondary myiasis may occur (Figure 2)

142 in some cases with complete or partial blindness (Figure 2). Blind animals may fall from

143 cliffs or die from starvation (28).

144 6. Diagnosis

145 The diagnosis of outbreaks of ocular disease in small ruminants is difficult,

146 especially in endemic areas. The disease must be differentiated from other similar clinic-

147 pathological syndromes such as PPR and pasteurellosis (53).

148 6.1.Traditional laboratory techniques

149 Mycoplasmas are fastidious organisms that are difficult to grow under laboratory

150 conditions (54). The gel precipitin test is a decades-old practice based on detection of

151 antigen (polysaccharide) in tissue specimens, particularly when specimens are no longer

152 suitable for culture because of deterioration during transit (55). The growth precipitation

153 test is for soluble cytoplasmic and extra-membranous antigens (56). The direct and indirect

154 fluorescent antibody tests are the most effective of the various serological methods

155 because of their simplicity and cost effectiveness (57). The latex agglutination test is

156 presently used routinely for outbreak investigation in Kenya (58). A competition ELISA

157 has also been developed (53), and has proved both specific and sensitive. However, these

158 methods are cumbersome, require specialized technical experience and reagents, and have

159 a high chance of cross-reaction or failure to identify the agent. Dark-field microscopy can
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160 be used for morphological studies (59). Biochemical tests are another option of diagnosis

161 but interspecies variation in some reactions should often need to take into consideration

162 (58).

163 6.2. Molecular techniques

164 M. conjunctivae may be better defined based on genomic features (60, 61). Using

165 the current taxonomic standards, online tools for the identification of Mycoplasma-related

166 bacteria have also been developed (62, 63). The species definition and identification are

167 based on general species definition using molecular techniques. A nucleic acid recognition

168 method has now been published, based on the amplification of various genes of the

169 mycoides cluster:16S rRNA (16), 23S rRNA (64) and 16S rDNA (65). These polymerase

170 chain reaction (PCR) techniques can be used directly on clinical materials (66). A serine-

171 rich membrane protein of M. conjunctivae, named LppS, was also characterized by gene

172 cloning and sequence analysis (28). TaqMan real-time PCR is considered a very rapid test

173 for M. conjuctivae genotyping in caprine (67-71).

174 7. Treatment and control

175 Generally, lipophilic drugs achieve higher intracorneal and intraocular

176 concentrations than hydrophilic drugs, and are more effective at penetrating the blood-tear

177 barrier. Trimethoprim and erythromycin are non-polar, basic, lipophilic drugs that

178 concentrate within tears following parenteral injection; however, they are not used

179 commercially because of poor activity against infection, the production of excessive tissue

180 inflammation or cost (72). The therapeutic opinions are summarized in Figure 3 and 4 and

181 discussed below.

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182 7.1. Systemic therapy

183 Oxytetracycline is an amphoteric molecule that should theoretically diffuse into

184 tears (73).Long-acting oxytetracycline (20 mg/kg, IM) is preferable, but treatment does

185 not assure complete effectiveness (59, 74). Florfenicol (20 mg/kg, I/M, S/C) has

186 significantly (p<0.05) higher tear concentration (75) and provides an effective treatment in

187 endemic regions where the use of oxytetracycline is restricted. Systemic non steroid anti-

188 inflammatory drugs (NSAIDs) may be used to provide relief from secondary uveitis (76).

189 In addition, autohemotherapy is effective in treatment of rapidly occurring corneal opacity

190 in caprines, possibly via animmune-stimulatory effect.

191 7.2. Topical therapy

192 Topical administration of antimicrobial formulations is seen as a potentially cost-

193 effective and less labour-intensive method of treatment (77). In Australia, Benzathine

194 cloxacillin ointment is registered for topical treatment f bovine cases, using a pre-

195 packed plastic application syringe (78). Tetracycline, oxytetracycline, polymyxin B or

196 erythromycin ointments are the drugs of choice, and should be applied 3–4 times daily for

197 4–7 days in small herds or flocks for which this is practical (79). Intramuscular injection

198 of oxytetracycline dehydrate, together with a topical application of chlortetracycline

199 hydrochloride powder, was highly effective in the treatment of affected ewes and lambs on

200 hill sheep (80). An eye-drop solution comprising 1% silver nitrate and 0.4% zinc sulphate

201 in sterile water was found effective in combating corneal opacity (81). If topical therapy is

202 not practical, addition of oxytetracycline to the feed (80mg/animal/day) may improve the

203 condition (82).

204 7.3. Sub–conjunctival therapy

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205 Goats with keratitis can be also treated daily with 0.1 mg of subconjunctival

206 injectable atropine to decrease ciliary spasm (83). Subconjunctival injection of

207 dexamethasone (0.5–1.0 mg/eye) is a procedure that is only undertaken with a few

208 animals (84) (Figure 4).

209 7.4. Therapeutic residue

210 After administration of a single dose of injectable oxytetracycline, the Food

211 Animal Residue Avoidance Databank (FARAD) recommends a discard period of at least

212 96 hours and then testing of milk for antibiotic residues (85). Following multiple doses or

213 high doses, a milk discard time of 144 hours should be observed, followed by residue

214 testing (85). The meat withdrawal interval is 5 days. After topical administration of

215 tetracyclines or tetracycline with polymyxin B, there is no required withdrawal time (86).

216 7.5. Additional management

217 Infected animals should be separated from the rest of the flock to prevent

218 mycoplasmal dissemination and outbreaks. Direct sunlight is irritating to the infected eyes

219 and should be avoided; it is best to keep the animals in a shady place such as in a stable or

220 under the tree. Because insects have a role in indirect mechanical transmission of IKC,

221 stables should be kept clean and dry, to discourage fly activity and create conditions that

222 are unfavourable for breeding. Natural or synthetic fly repellent could be used in fly hot

223 spots, and there is growing interest in the use of fly-repellent insecticide treated nets. For

224 animals severely affected with IKC, provision of temporary tarsorrhaphy or third eyelid

225 flap may give additional protection from the environment and prevent exposure in globes

226 with a markedly distended cornea (87). Finally, animals should be given adequate feed and

227 water, with care and assistance, as they may unable to graze.
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228 8. Future antimicrobial research to combat M. conjunctivae

229 Effectiveness of antimicrobials to cure IKC has less studied than other

230 mycoplasmal diseases of livestock and poultry. The activities of danofloxacin and tylosin

231 (macrolide) found great potential for the treatment of respiratory and other infections

232 caused by Mycoplasma species in cattle, pigs and poultry (88). Enrofloxacin,

233 ciprofloxacin, tylosin, and doxycycline are the most effective antimicrobials in vitro, in

234 comparable to, tetracycline, lincomycin: spectinomycin and spiramycin against field

235 isolates of Mycoplasma agalactiae (89). In Japan, Mycoplasma species (M. bovirhinis, M.

236 bovis, M. alkalescens and M. bovoculi) isolated from homebred cattle was susceptible to

237 spectinomycin and fluoroquinolones but resistant to macrolides, tetracyclines,

238 thiamphenicol and flumequine, whereas kanamycin, lincomycin and chloramphenicol

239 maintained intermediate effectiveness against these species (90, 91). However increase in

240 resistance to erythromycin, tilmicosin, tylosin, ciprofloxacin, enrofloxacin,

241 chlortetracycline, doxycycline, and oxytetracycline are also a major concerned (92).

242 It is hard to reach a conclusion to choose an appropriate antimicrobial against IKC without

243 sensitivity testing and field trial. Future research should be conducted with every

244 antimicrobial effective against other mycoplasmal diseases to draw a standard outcome in

245 treatment of IKC.

246 9. Economic impact

247 Small ruminants are important commodities to much of the world's population as a

248 source of meat, milk and skin. IKC is a sporadic disease of economic importance globally,

249 posing a major constraint to goat production. Direct losses from the disease result from its

250 high rate of blindness, reduced milk and meat yield, cost of treatment, control, disease
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251 diagnosis and surveillance. In addition, there are indirect losses due to the imposition of

252 trade restrictions in the marketing channel.

253 10. Welfare issues

254 Clearly, there are welfare issues with small ruminants with IKC, and their

255 discomfort from eye disease should be taken seriously, as it would in a human being (93).

256 IKC often results in a dull, depressed animal that has a reduced appetite and body

257 condition (94). These factors have negative effects on both animal welfare and production.

258 The condition appears to be more serious in the wild, where there is no chance of human

259 assistance and the animal succumbs to prey or dies through starvation.

260 11. People react towards infectious keratoconjunctivitis

261 Those tending livestock become anxious about an animal when it suffers from IKC.

262 It is hard to detect an eye problem in a single animal in a flock, but quite easy to spot in an

263 individual animal or household rearing system, which is common in urban areas.

264 Occasionally, owners try to resolve the eye problem using antibiotic eye drops available in

265 the market for human use. More often, farmers keep their animal at home, based on

266 thinking that the problem will resolve after 2–3 days, as uncomplicated seasonal

267 conjunctivitis does in human. However, this malpractice makes the IKC worse, due to

268 development of ulcer and corneal opacity. Owners worry about the eyes of their livestock,

269 because animals without vision have poor market value, or the owner may believe they

270 will seem pathetic in choosing to rear a blind animal. However, owners often have more

271 urgency in treating a pregnant animal, to give the offspring a better chance of survival.

272 12. Prevention

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273 There is no known vaccination available to provide long-lasting immunity against

274 IKC. Therefore, biosecurity measures are the easiest option. Quarantine of incoming

275 animals for at least 2 weeks maybe beneficial, but is not often practical for control of the

276 disease (59). The provision of adequate space in houses and particularly at feed troughs is

277 also recommended (34).

278 13. Conclusion

279 IKC can cause a severe widespread outbreak of blindness in small ruminants. A

280 complete ocular examination is critical when animals present with an ocular complaint,

281 and early identification of the disease is of paramount importance in reducing or

282 preventing the spread of infection. Epidemiological and clinical research is needed to

283 establish risk factors for this disease, especially in the tropics, where cases are found

284 sporadically but few studies have been undertaken. A better understanding of the effect of

285 IKC on animal health, welfare and production is needed. At the same time, the

286 effectiveness of macrolide or other group of antimicrobials should also be included in field

287 research.

288 Acknowledgement

289 The author is grateful to the owners and the students who shared their experiences

290 with me during the case study and therapeutic periods. I would like to thank Dr Hilary

291 Cadman, PhD (Cadman Editing Services NSW, Australia) for providing English-language

292 editing of this article.

293 Conflicts of interest

294 The author has nothing to disclose.

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295 References

296 1. Lysnyansky, I., Levisohn, S., Bernstein, M., Kenigswald, J., Blum, S., Gerchman, I., Elad, D.,
297 and Brennee, J. 2007. Diagnosis of Mycoplasma conjunctivae and Chlamydophila spp in
298 an episode of conjunctivitis/keratoconjunctivitis in lambs. Israel Journal of Veterinary
299 Medicine 62:79-82.
300 2. Akerstedt, J., and Hofshagen, M. 2004. Bacteriological investigation of infectious
301 keratoconjunctivitis in Norwegian sheep. Acta Veterinaria Scandinavica 45:19-26.
302 3. Scott, P.R. 2015. Sheep medicine: CRC Press.
303 4. Chakrabarti, A., Kumar, P., Chandran, P.C., Dey, A., and Dayal, S. 2014. Prevalence of Eye
304 Diseases of Cattle in Bihar, India. Journal of Animal Health and Production 2:25-27.
305 5. Egwu, G., Faull, W., Bradbury, J., and Clarkson, M. 1989. Ovine infectious
306 keratoconjunctivitis: a microbiological study of clinically unaffected and affected sheep's
307 eyes with special reference to Mycoplasma conjunctivae. The Veterinary record 125:253-
308 256.
309 6. Loria, G., Puleio, R., Agnello, S., Chetta, M., Tola, S., and Nicholas, R. 2011. Infectious
310 kerato-conjunctivitis (IKC) caused by Mycoplasma conjunctivae: a new risk for Italian
311 sheep farming. Large Animal Review 17:13-17.
312 7. Yeruham, I., Perl, S., Hadani, A., Elad, D., and De-Lange, A. 1991. Survey of eye pathology
313 in a dairy herd in the Jericho Valley in Israel. Israel Journal of Veterinary Medicine 46:142-
314 144.
315 8. Osman, K., Ali, H., ElJakee, J., and Galal, H. 2011. Chlamydophila psittaci and
316 Chlamydophila pecorum infections in goats and sheep in Egypt. Revue scientifique et
317 technique (International Office of Epizootics) 30:939-948.
318 9. Bankemper, K.W., Lindley, D.M., Nusbaum, K.E., and Mysinger, R.H. 1990.
319 Keratoconjunctivitis associated with Neisseria ovis infection in a herd of goats. J. Vet.
320 Diagn. Investig 2:76-78.
321 10. Langford, E. 1971. Mycoplasma and associated bacteria isolated from ovine pink-eye.
322 Canadian Journal of Comparative Medicine 35:18.
323 11. Kummeneje, K., and Mikkelsen, T. 1975. Isolation of Listeria monocytogenes type 04 from
324 cases of keratoconjunctivitis in cattle and sheep. Nordisk veterinaermedicin 27:144-149.
325 12. Abbas, B., Razig, S., and Shigidi, M. 1979. The aetiology of a keratoconjunctivitis occurring
326 in goats in the Sudan. The Veterinary record 105:348-350.
327 13. Moore, C., and Whitley, R. 1984. Ophthalmic diseases of small domestic ruminants. The
328 Veterinary clinics of North America. Large animal practice 6:641.
329 14. Van Halderen, A., Van Rensburg, W., Geyer, A., and Vorster, J. 1994. The identification of
330 Mycoplasma conjunctivae as an aetiological agent of infectious keratoconjunctivitis of
331 sheep in South Africa. The Onderstepoort journal of veterinary research 61:231-237.
332 15. Trotter, S., Franklin, R., Baas, E., and Barile, M. 1977. Epidemic caprine
333 keratoconjunctivitis: experimentally induced disease with a pure culture of Mycoplasma
334 conjunctivae. Infection and immunity 18:816-822.
335 16. Giacometti, M., Nicolet, J., Frey, J., Krawinkler, M., Meier, W., Welle, M., Johansson, K.-E.,
336 and Degiorgis, M.-P. 1998. Susceptibility of alpine ibex to conjunctivitis caused by
337 inoculation of a sheep-strain of Mycoplasma conjunctivae. Veterinary microbiology
338 61:279-288.
339 17. Degiorgis, M., Frey, J., Nicolet, J., Abdo, E.-M., Fatzer, R., Schlatter, Y., Reist, S., Janovsky,
340 M., and Giacometti, M. 2000. An outbreak of infectious keratoconjunctivitis in Alpine
341 chamois (Rupicapra r. rupicapra) in Simmental-Gruyères, Switzerland. SAT, Schweizer
342 Archiv für Tierheilkunde 142:520-527.
343 18. Baker, S., Bashiruddin, J., Ayling, R., and Nicholas, R. 2001. Molecular detection of
 Page 15 of 23

344 Mycoplasma conjunctivae in English sheep affected by infectious keratoconjunctivitis.

345 Veterinary Record 148:240-241.
346 19. Pettersson, B., Uhlen, M., and Johansson, K.-E. 1996. Phylogeny of some mycoplasmas
347 from ruminants based on 16S rRNA sequences and definition of a new cluster within the
348 hominis group. International journal of systematic bacteriology 46:1093-1098.
349 20. Barile, M.F., Del Giudice, R.A., and Tully, J.G. 1972. Isolation and characterization of
350 Mycoplasma conjunctivae sp. n. from sheep and goats with keratoconjunctivitis. Infection
351 and immunity 5:70-76.
352 21. Ter Laak, E., Schreuder, B., and Smith‐Buys, C. 1988. The occurrence of Mycoplasma
353 conjunctivae in the Netherlands and its association with infectious keratoconjunctivitis in
354 sheep and goats. Veterinary Quarterly 10:73-83.
355 22. Jansen, B.D., Heffelfinger, J.R., Noon, T.H., Krausman, P.R., and deVos Jr, J.C. 2006.
356 Infectious keratoconjunctivitis in bighorn sheep, Silver Bell Mountains, Arizona, USA.
357 Journal of Wildlife Diseases 42:407-411.
358 23. Marco, I., Mentaberre, G., Ballesteros, C., Bischof, D.F., Lavín, S., and Vilei, E.M. 2009.
359 First report of Mycoplasma conjunctivae from wild Caprinae with infectious
360 keratoconjunctivitis in the Pyrenees (NE Spain). Journal of wildlife diseases 45:238-241.
361 24. Evans, A.L., Bey, R.F., Schoster, J.V., Gaarder, J.E., and Finstad, G.L. 2008. Preliminary
362 studies on the etiology of keratoconjunctivitis in reindeer (Rangifer tarandus tarandus)
363 calves in Alaska. Journal of wildlife diseases 44:1051-1055.
364 25. Høvding, G. 2008. Acute bacterial conjunctivitis. Acta ophthalmologica 86:5-17.
365 26. Bielory, L., Lyons, K., and Goldberg, R. 2012. Climate change and allergic disease. Current
366 allergy and asthma reports 12:485-494.
367 27. Gouws, J., Coetzer, J., and Howell, P. 1995. A comparative microbiological study of
368 clinically healthy eyes and those affected by ophthalmia in cattle and the association of
369 noctuid eye-frequenting moths. Journal of the South African Veterinary Association
370 66:160-169.
371 28. Belloy, L., Janovsky, M., Vilei, E.M., Pilo, P., Giacometti, M., and Frey, J. 2003. Molecular
372 epidemiology of Mycoplasma conjunctivae in Caprinae: transmission across species in
373 natural outbreaks. Applied and environmental microbiology 69:1913-1919.
374 29. Janovsky, M., Frey, J., Nicolet, J., Belloy, L., Goldschmidt-Clermont, E., and Giacometti, M.
375 2001. Mycoplasma conjunctivae infection is self-maintained in the Swiss domestic sheep
376 population. Veterinary microbiology 83:11-22.
377 30. Jimena, O.N., Laura, J.M., Elena, M.M.R., Alonso, N.H.J., and Teresa, Q.M.M. 2009.
378 Association of Raillietia caprae with the presence of Mycoplasmas in the external ear
379 canal of goats. Preventive veterinary medicine 92:150-153.
380 31. DaMassa, A., and Brooks, D. 1991. The external ear canal of goats and other animals as a
381 mycoplasma habitat. Small Ruminant Research 4:85-93.
382 32. Šeol, B., Hajsig, D., Busch, K., Lojkić, M., and Naglić, T. 2000. Epidemiological and
383 microbiological study of an outbreak of infectious keratoconjunctivitis in sheep.
384 Veterinary Record 147:72-75.
385 33. Postma, G.C., Carfagnini, J.C., and Minatel, L. 2008. Moraxella bovis pathogenicity: an
386 update. Comparative immunology, microbiology and infectious diseases 31:449-458.
387 34. Kováč, G., Nagy, O., Seidel, H., Pilipčinec, E., Novotný, J., and Link, R. 2003. Infectious
388 keratoconjunctivitis in sheep. VALOCKÝ, I., KRAJNIČÁKOVÁ, M., MOJŽIŠOVÁ, J.,
389 BUGARSKÝ, A., FIALKOVIČOVÁ, M. 47:130-134.
390 35. Hadani, Y., Lysnyansky, I., Bareli, A., Bernstein, M., Elad, D., Ozeri, R., Rotenberg, D.,
391 Bumbarov, V., Perl, S., and Brenner, J. 2013. An Unusual Widespread Outbreak of
392 Blindness Caused by Mycoplasma conjunctivae on a Large Dairy Goat Farm. Front Cover-
393 THE SPHYNX BREED OF CAT 68:3.
394 36. Snowder, G., Van Vleck, L.D., Cundiff, L., and Bennett, G. 2005. Genetic and
 Page 16 of 23

395 environmental factors associated with incidence of infectious bovine keratoconjunctivitis

396 in preweaned beef calves. Journal of animal science 83:507-518.
397 37. Hosie, B. 1989. Infectious keratoconjunctivitis in sheep and goats. Veterinary annual.
398 38. Mavrot, F., Zimmermann, F., Vilei, E.M., and Ryser-Degiorgis, M.-P. 2012. Is the
399 development of infectious keratoconjunctivitis in Alpine ibex and Alpine chamois
400 influenced by topographic features? European journal of wildlife research 58:869-874.
401 39. Zielinski, G., Young, T., Ross, R., and Rosenbusch, R. 1990. Adherence of Mycoplasma
402 hyopneumoniae to cell monolayers. American journal of veterinary research 51:339-343.
403 40. Zielinski, G., and Ross, R. 1993. Adherence of Mycoplasma hyopneumoniae to porcine
404 ciliated respiratory tract cells. American journal of veterinary research 54:1262-1269.
405 41. Minion, F.C., Adams, C., and Hsu, T. 2000. R1 region of P97 mediates adherence
406 ofmycoplasma hyopneumoniae to swine cilia. Infection and immunity 68:3056-3060.
407 42. Burnett, T.A., Dinkla, K., Rohde, M., Chhatwal, G.S., Uphoff, C., Srivastava, M., Cordwell,
408 S.J., Geary, S., Liao, X., and Minion, F.C. 2006. P159 is a proteolytically processed, surface
409 adhesin of Mycoplasma hyopneumoniae: defined domains of P159 bind heparin and
410 promote adherence to eukaryote cells. Molecular microbiology 60:669-686.
411 43. Pilo, P., Vilei, E.M., Peterhans, E., Bonvin-Klotz, L., Stoffel, M.H., Dobbelaere, D., and Frey,
412 J. 2005. A metabolic enzyme as a primary virulence factor of Mycoplasma mycoides
413 subsp. mycoides small colony. Journal of bacteriology 187:6824-6831.
414 44. Bischof, D.F., Janis, C., Vilei, E.M., Bertoni, G., and Frey, J. 2008. Cytotoxicity of
415 Mycoplasma mycoides subsp. mycoides small colony type to bovine epithelial cells.
416 Infection and immunity 76:263-269.
417 45. Zimmermann, L., Peterhans, E., and Frey, J. 2010. RGD motif of lipoprotein T, involved in
418 adhesion of Mycoplasma conjunctivae to lamb synovial tissue cells. Journal of
419 bacteriology 192:3773-3779.
420 46. Pierschbacher, M.D., and Ruoslahti, E. 1983. Cell attachment activity of fibronectin can be
421 duplicated by small synthetic fragments of the molecule. Nature 309:30-33.
422 47. Ruoslahti, E. 2003. The RGD story: a personal account. Matrix Biology 22:459-465.
423 48. Calderon-Copete, S.P., Wigger, G., Wunderlin, C., Schmidheini, T., Frey, J., Quail, M.A.,
424 and Falquet, L. 2009. The Mycoplasma conjunctivae genome sequencing, annotation and
425 analysis. BMC bioinformatics 10:S7.
426 49. Tryland, M., Neves, C.G.D., Sunde, M., and Mørk, T. 2009. Cervid herpesvirus 2, the
427 primary agent in an outbreak of infectious keratoconjunctivitis in semidomesticated
428 reindeer. Journal of clinical microbiology 47:3707-3713.
429 50. Rehbinder, C., and Nilsson, A. 1995. An outbreak of kerato-conjunctivitis among corralled,
430 supplementary fed, semi-domestic reindeer calves. Rangifer 15:9-14.
431 51. El-Tookhy, O., and Tharwat, M. 2013. Clinical and ultrasonographic findings of some
432 ocular conditions in sheep and goats. Open Veterinary Journal 3:11-16.
433 52. Shahzad, W., Munir, R., Rana, M.Y., Ahmad, R., Khan, M.S., Akbar, G., Ijaz, M., and
434 Mehmood, F. 2013. Prevalence, molecular diagnosis and treatment of Mycoplasma
435 conjunctivae isolated from infectious keratoconjunctivitis affected Lohi sheep maintained
436 at Livestock Experiment Station, Bahadurnagar, Okara, Pakistan. Tropical animal health
437 and production 45:737-742.
438 53. Thiaucourt, F., and Bölske, G. 1996. Contagious caprine pleuropneumonia and other
439 pulmonary mycoplasmoses of sheep and goats. Revue scientifique et technique
440 (International Office of Epizootics) 15:1397-1414.
441 54. Citti, C., and Blanchard, A. 2013. Mycoplasmas and their host: emerging and re-emerging
442 minimal pathogens. Trends in microbiology 21:196-203.
443 55. Rurangirwa, F., McGuire, T., Kibor, A., and Chema, S. 1987. A LATEX AGGLUTINATION TEST
444 FOR FIELD DIAGONOIS OF CONTAGIOUS CAPRINE PHEUROPNEUMONIA. SMALL
445 RUMINANT:82.
 Page 17 of 23

446 56. Krogsgaard-Jensen, A. 1972. Mycoplasma: growth precipitation as a serodiagnostic

447 method. Applied microbiology 23:553-558.
448 57. Rosendal, S., and Black, F. 1972. Direct and indirect immunofluorescence of unfixed and
449 fixed mycoplasma colonies. Acta Pathologica Microbiologica Scandinavica Section B
450 Microbiology and Immunology 80:615-622.
451 58. Houshaymi, B., Tekleghiorghis, T., Wilsmore, A., Miles, R., and Nicholas, R. 2002.
452 Investigations of outbreaks of contagious caprine pleuropneumonia in Eritrea. Tropical
453 animal health and production 34:383-389.
454 59. Nicholas, R., Ayling, R., and Loria, G. 2008. Ovine mycoplasmal infections. Small Ruminant
455 Research 76:92-98.
456 60. McAuliffe, L., Ellis, R.J., Lawes, J.R., Ayling, R.D., and Nicholas, R.A. 2005. 16S rDNA PCR
457 and denaturing gradient gel electrophoresis; a single generic test for detecting and
458 differentiating Mycoplasma species. Journal of medical microbiology 54:731-739.
459 61. Thompson, C.C., Vieira, N.M., Vicente, A.C.P., and Thompson, F.L. 2011. Towards a
460 genome based taxonomy of Mycoplasmas. Infection, Genetics and Evolution 11:1798-
461 1804.
462 62. Stackebrandt, E., Frederiksen, W., Garrity, G.M., Grimont, P.A., Kämpfer, P., Maiden,
463 M.C., Nesme, X., Rosselló-Mora, R., Swings, J., and Trüper, H.G. 2002. Report of the ad
464 hoc committee for the re-evaluation of the species definition in bacteriology.
465 International journal of systematic and evolutionary microbiology 52:1043-1047.
466 63. Zhao, Y., Wei, W., Lee, M., Shao, J., Suo, X., and Davis, R.E. 2009. Construction of an
467 interactive online phytoplasma classification tool, iPhyClassifier, and its application in
468 analysis of the peach X-disease phytoplasma group (16SrIII). International Journal of
469 Systematic and Evolutionary Microbiology 59:2582-2593.
470 64. Schnee, C., Schulsse, S., Hotzel, H., Ayling, R.D., Nicholas, R.A., Schubert, E., Heller, M.,
471 Ehricht, R., and Sachse, K. 2012. A novel rapid DNA microarray assay enables
472 identification of 37 Mycoplasma species and highlights multiple Mycoplasma infections.
473 65. Taghavi Razavizadeh, A., and Razmyar, J. 2014. Molecular diagnosis of Mycoplasma
474 conjunctivae in an outbreak of infectious keratoconjunctivitis in sheep. Iranian Journal of
475 Veterinary Research 15:72-74.
476 66. Bölske, G., Mattsson, J.G., Bascuñana, C.R., Bergström, K., Wesonga, H., and Johansson,
477 K.-E. 1996. Diagnosis of contagious caprine pleuropneumonia by detection and
478 identification of Mycoplasma capricolum subsp. capripneumoniae by PCR and restriction
479 enzyme analysis. Journal of clinical microbiology 34:785-791.
480 67. Vilei, E.M., Bonvin-Klotz, L., Zimmermann, L., Ryser-Degiorgis, M.-P., Giacometti, M., and
481 Frey, J. 2007. Validation and diagnostic efficacy of a TaqMan real-time PCR for the
482 detection of Mycoplasma conjunctivae in the eyes of infected Caprinae. Journal of
483 microbiological methods 70:384-386.
484 68. Ryser-Degiorgis, M.-P., Bischof, D.F., Marreros, N., Willisch, C., Signer, C., Filli, F., Brosi, G.,
485 Frey, J., and Vilei, E.M. 2009. Detection of Mycoplasma conjunctivae in the eyes of
486 healthy, free-ranging Alpine ibex: possible involvement of Alpine ibex as carriers for the
487 main causing agent of infectious keratoconjunctivitis in wild Caprinae. Veterinary
488 microbiology 134:368-374.
489 69. Mavrot, F., Vilei, E.M., Marreros, N., Signer, C., Frey, J., and Ryser-Degiorgis, M.-P. 2012.
490 Occurrence, quantification, and genotyping of Mycoplasma conjunctivae in wild Caprinae
491 with and without infectious keratoconjunctivitis. Journal of wildlife diseases 48:619-631.
492 70. Arnal, M., Herrero, J., de la Fe, C., Revilla, M., Prada, C., Martínez-Durán, D., Gómez-
493 Martín, Á., Fernández-Arberas, O., Amores, J., and Contreras, A. 2013. Dynamics of an
494 infectious keratoconjunctivitis outbreak by Mycoplasma conjunctivae on pyrenean
495 chamois rupicapra p. Pyrenaica. PloS one 8.
496 71. Fernández-Aguilar, X., Cabezón, Ó., Marco, I., Mentaberre, G., Frey, J., Lavín, S., and
 Page 18 of 23

497 López-Olvera, J.R. 2013. Mycoplasma conjunctivae in domestic small ruminants from high
498 mountain habitats in Northern Spain. BMC veterinary research 9:253.
499 72. Dueger, E., Angelos, J., Cosgrove, S., Johnson, J., and George, L. 1999. Efficacy of
500 florfenicol in the treatment of experimentally induced infectious bovine
501 keratoconjunctivitis. American journal of veterinary research 60:960-964.
502 73. Punch, P., Costa, N., Chambers, E., Slatter, D., and Wilcox, G. 1985. Plasma and tear
503 concentrations of antibiotics administered parenterally to cattle. Research in veterinary
504 science 39:179-187.
505 74. Hosie, B., and Greig, A. 1995. Role of oxytetracycline dihydrate in the treatment of
506 mycoplasma associated ovine keratoconjunctivitis in lambs. British Veterinary Journal
507 151:83-88.
508 75. Regnier, A., Laroute, V., Gautier-Bouchardon, A., Gayrard, V., Picard-Hagen, N., and
509 Toutain, P.-L. 2013. Florfenicol concentrations in ovine tear fluid following intramuscular
510 and subcutaneous administration and comparison with the minimum inhibitory
511 concentrations against mycoplasmal strains potentially involved in infectious
512 keratoconjunctivitis. American journal of veterinary research 74:268-274.
513 76. Angelos, J.A. 2013. Overview of Infectious Keratoconjunctivitis. In The Merck Veterinary
514 Manual S.L. Cynthia M. Kahn, editor: The Merck Publishing Group.
515 77. O'Connor, A.M., Wellman, N.G., Evans, R.B., and Roth, D.R. 2006. A review of randomized
516 clinical trials reporting antibiotic treatment of infectious bovine keratoconjunctivitis in
517 cattle. Animal Health Research Reviews 7:119-127.
518 78. Daigneault, J., George, L., and Baggot, J. 1990. Ocular and serum disposition kinetics of
519 cloxacillin after topical administration of benzathine cloxacillin and intravenous
520 administration of sodium cloxacillin to calves. American journal of veterinary research
521 51:381-385.
522 79. Ward, D., and Clark, E. 1991. Ocular pharmacology. The Veterinary clinics of North
523 America. Food animal practice 7:779-791.
524 80. Hosie, B. 1988. Keratoconjunctivitis in a hill sheep flock. The Veterinary record 122:40-43.
525 81. Jeyabal, L., Ray, D., Sureshkannan, S., Nagarajan, K., Visnuvinayagam, S., Ghosh, S.,
526 Banerjee, P.S., Sekar, S.C., Bagath, M., and Padmanath, K. 2013. First Report of Moraxella
527 Bovis Infection in Indian Cattle.
528 82. Kahn, C.M., Line, S., and Allen, D.G. 2005. The Merck veterinary manual: Merck
529 Whitehouse Station, NJ.
530 83. Peiffer, R., and Stowe, C. 1981. Veterinary ophthalmic pharmacology. Veterinary
531 Ophthalmology. 2nd ed. Philadelphia: Lea & Febiger:160-205.
532 84. Alexander, D. 2010. Infectious bovine keratoconjunctivitis: a review of cases in clinical
533 practice. Veterinary Clinics of North America: Food Animal Practice 26:487-503.
534 85. Martin-Jimenez, T., Craigmill, A., and Riviere, J. 1997. Extralabel use of oxytetracycline.
535 Journal of the American Veterinary Medical Association (USA).
536 86. Webb, A.I., Baynes, R.E., Craigmill, A.L., Riviere, J.E., and Haskell, S.R. 2004. FARAD Digest.
537 87. Townsend, W.M. 2010. Examination techniques and therapeutic regimens for the
538 ruminant and camelid eye. Veterinary Clinics of North America: Food Animal Practice
539 26:437-458.
540 88. Cooper, A., Fuller, J., Fuller, M., Whittlestone, P., and Wise, D. 1993. In vitro activity of
541 danofloxacin, tylosin and oxytetracycline against mycoplasmas of veterinary importance.
542 Research in veterinary science 54:329-334.
543 89. Loria, G., Sammartino, C., Nicholas, R., and Ayling, R. 2003. In vitro susceptibilities of field
544 isolates of Mycoplasma agalactiae to oxytetracycline, tylosin, enrofloxacin, spiramycin
545 and lincomycin–spectinomycin. Research in Veterinary Science 75:3-7.
546 90. Uemura, R., Sueyoshi, M., and Nagatomo, H. 2010. Antimicrobial susceptibilities of four
547 species of Mycoplasma isolated in 2008 and 2009 from cattle in Japan. Journal of
 Page 19 of 23

548 Veterinary Medical Science 72:1661-1663.

549 91. Kawai, K., Higuchi, H., Iwano, H., Iwakuma, A., Onda, K., Sato, R., Hayashi, T., Nagahata,
550 H., and Oshida, T. 2014. Antimicrobial susceptibilities of Mycoplasma isolated from
551 bovine mastitis in Japan. Animal Science Journal 85:96-99.
552 92. Gharaibeh, S., and Al-Rashdan, M. 2011. Change in antimicrobial susceptibility of
553 Mycoplasma gallisepticum field isolates. Veterinary microbiology 150:379-383.
554 93. Webster, A. 2001. Farm animal welfare: the five freedoms and the free market. The
555 veterinary journal 161:229-237.
556 94. Williams, D.L. 2010. Welfare issues in farm animal ophthalmology. Veterinary Clinics of
557 North America: Food Animal Practice 26:427-435.
558 95. Vemuganti, G.K., Murthy, S.I., and Das, S. 2011. Update on pathologic diagnosis of corneal
559 infections and inflammations. Middle East African journal of ophthalmology 18:277.

560

561 List of picture

Figure 1. Two years old domesticated Black Bengal Goat is suffering from IKC since three
days with the signs of lacrimation, hyperemia and corneal opacity admitted at Teaching
Veterinary Hospital (TVH) of Chittagong Veterinary & Animal Sciences University
(CVASU), Bangladesh.

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Figure 2. The eye of an infectious keratoconjunctivitis affected Jamunapari goat becomes

secondarily infested with maggots of fly (opthalmomyiasis) due to ignorance of owner to
treat in time and proper care. (Location: TVH of CVASU, Bangladesh)

568
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Subconjunctival Intramuscular Intramuscular

Figure 3. Combined treatment approach of Infectious keratoconjunctivitis using

subconjunctival dexamethasone sodium (0.2-0.4 mg/eye), Autohemotherapy (3-5 ml
IM/goat/week) and long acting oxytetracycline (20mg/kg body weight IM/48 hours
interval. This protocol was applied at TVH of CVASU.

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Figure 4. Sub-conjunctival administration of dexamethasone @0.5 mg/eye using insulin

syringe in an IKC infected goat patient at TVH of CVASU, Bangladesh.

576

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578

Table 1: Summary of histological features in infection in cornea (95)

Anatomic Histologic features to be observed

layer
General Thickness, thinning, necrosis, perforation, separation, exudates,
features pigmentation

Epithelium Intactness, edema, ulceration, hyperplasia, down growths

inflammatory infiltrates, giant cell reaction, cytoplasmic
inclusions regularity/breaks of basement membrane, pannus
formation(inflammatory or degenerative)

Bowman’s Thickness, breaks, absence, calcification, degenerative changes,

 Page 23 of 23

layer any deposits

Stroma Thinning, edema, vascularization, inflammation and density and

type of inflammatory cells (neutrophils, lymphocytes, plasma
cells, giant cells, location of cells (anterior/ mid/posterior
stroma), perforation, cellularity, changes in keratocytes
(myofibroblastic transformation, loss of keratocytes) orientation
of collagen fibers, fibrosis, scarring, abnormal deposits, any
infectious agent and its load and location

Descemet’s Thin, fragmented or intact. Giant cell reaction around DM,

membrane granulomatous inflammation around fragmented ends
(DM) and Presence of microorganisms, presence and adequacy of
endothelium endothelial cells, morphology of endothelial cells, retrocorneal
membrane and exudates adherent to DM, anterior chamber
exudates

Others Adherent uveal tissue, AC exudates

579

580