Skip to main content

Low prevalence of Moraxella catarrhalis in the patients who suffered from conjunctivitis in the southwest of Iran

Abstract

Objective

Moraxella catarrhalis is a non-motile Gram-negative diplococcus bacterium that contributed to several human infections including conjunctivitis. This study aimed to reveal the prevalence of M. catarrhalis in patients who suffered from conjunctivitis in Ahvaz city, southwest of Iran.

Results

Out of 100 conjunctiva swab specimens, M. catarrhalis was isolated only from one (1%) conjunctivitis cases using the culture method. This strain was isolated from a 34 years old female patient. Also, the results of the polymerase chain reaction (PCR) were in agreement with the culture method, and the specimen that showed positive culture was also positive for specific gene of M. catarrhalis. The remaining 99 specimens did not show positive results with any of the culture and PCR methods.

Introduction

Moraxella catarrhalis is a Gram-negative, diplococcus bacterium that accounts for many pathologies of humans including otitis media, sinusitis, pneumonia, and conjunctivitis [1,2,3]. The inflammation of the membrane of eyelids is called conjunctivitis [4]. Several numbers of infectious pathogens including bacteria (Staphylococcus aureus, Haemophilus influenzae, Streptococcus pneumoniae, and Moraxella catarrhalis) and viruses (adenoviruses, herpes simplex virus, varicella-zoster virus, and Molluscum contagiosum) are contributed to the majority of conjunctivitis cases [5]. The incidence of viral conjunctivitis is higher than bacterial conjunctivitis in adults, whereas bacterial conjunctivitis is more common in children [6].

In total, bacterial conjunctivitis is accountable for 50–75% of cases in children [6]. Staphylococcus spp, S. pneumoniae, and H. influenzae account for the highest causes of bacterial conjunctivitis [5, 6]. In previous studies, the non-typeable H. influenzae (NTHi) had a frequency rate of 61.8% in children with infective conjunctivitis, followed by S. pneumoniae (28.2%), and M. catarrhalis (19.1%) [7]. In the last century, M. catarrhalis has been considered an emerging human pathogen [8]. This bacterium is resistant to penicillin due to its BRO-1 and BRO-2 beta-lactamases [8]. Conjunctivitis caused by M. catarrhalis is generally non-exudative, persistent, and with no sign of redness [9]. Currently, medical system laboratories do not have an identical sensitive method for differentiating Moraxella species to genus and species levels in eye infections [3]. Although the prevalence of Moraxella species in ocular infections has been rarely reported (0.8–19.1%) [3, 10], since there was no epidemiological information in this field in the geographical region of southwestern Iran, this study aimed to investigate the phenotypic and molecular presence of M. catarrhalis in conjunctivitis samples collected from patients admitted to one of the main referral ocular infections centers of southwestern Iran.

Main text

Methods

Sample collection and phenotypic detection of bacteria

In this study, which was performed from May 2013 to August 2016, the conjunctival samples were collected by sterile cotton-wool swabs (2 swabs) from each patient who suffered from infective conjunctivitis and admitted as outpatients to the Ophthalmology Division of Emam Khomeini Hospital in Ahvaz, southwestern Iran. The cases were selected and confirmed by the ophthalmologist attending physician. All cases with a history of antibiotics use 15 days before the study were excluded from research. The swab specimens were directly inoculated on blood agar (Merck, Darmstadt, Germany) with 5% defibrinated sheep blood (Bahar Afshan, Tehran, Iran) and incubated at 37 °C with 5% CO2 for 24–48 h. After overnight incubation, positive growth and suspected colonies were further investigated by colony morphology (smooth, white, opaque), Gram staining, oxidase, catalase, DNase, reduction of nitrate to nitrite, and carbohydrate fermentation tests such as sucrose, glucose, and lactose [11]. The suspected colonies were stocked in trypticase soy broth (Merck, Germany) containing 20% (v/v) glycerol and frozen at − 80 °C. The isolates were finally confirmed by polymerase chain reaction (PCR) assay.

Molecular investigation

DNA extraction and PCR assay

The DNA extraction of bacterial isolates and conjunctival swabs was done by the boiling method and QIAamp DNA Mini Kit (QIAGEN GmbH, Hilden, Germany), respectively [12,13,14]. The PCR was done to amplify a 140 bp sequence of M. catarrhalis using MCAT1 (5′-TTGGCTTGTGCT AAAATATC-3′) and MCAT2 (5′-GTCATCGCTATCATTCACCT-3′) primers as previously described by Post et al. [15]. The PCR reaction was carried out in a final volume of 25 µl using a thermal gradient cycler (Eppendorf Co., Germany) with the following procedure: 2.5 µl of 10× buffer (10 mM Tris-HCl, 50 mM KCl), 1.5 mM MgCl2, 200 µM of each dNTPs, 0.4 µM of each forward and reverse primers, 1 U of Taq polymerase, 3 µl of the DNA template, and sterilized distilled water to complete the reaction volume. The PCR program was optimized as initial denaturation at 95 °C for 5 min, 34 cycles of denaturation at 95 °C for 50s, annealing at 55 °C for 50s, extension at 72 °C for 50s, and the final extension time at 72 °C for 5 min. The PCR amplicons were visualized by electrophoresis (80 V/60 min) on 1.5% agarose gel in 1× Tris Borate-EDTA buffer containing ethidium bromide (0.5 µg/ml). M. catarrhalis ATCC 25238 and distilled water were used as positive and negative controls respectively.

Results

In this study, a total of 100 patients were enrolled from which 65 and 35 were males and females, respectively. The mean age was 45 years (range 7–76 years). The result of the culture of conjunctival swab specimens showed that only one sample was positive for M. catarrhalis growth. This isolate had a positive reaction for catalase, oxidase, nitrate reduction, and DNase (Table 1); also exhibited a positive band of 140 bp in PCR assay (Fig. 1). M. catarrhalis was isolated from a 34 year old female patient. The other 99 specimens were negative for M. catarrhalis by culture method. Also, the PCR method detected the M. catarrhalis in only one of the extracted DNA samples that belong to the culture positive specimen. The remaining 99 specimens were negative for the specific band of M. catarrhalis by PCR.

Table 1. Biochemical characteristics of Moraxella catarrhalis isolated from conjunctival swab.
Fig. 1
figure1

Lanes 1: ladder of 50 bp; lane 2: positive control, lane 3: negative control, lane 4: the amplicon of bacterial isolate, lane 5: the amplicon of conjunctival sample

Discussion

Conjunctivitis is one of the main causes of redness and discharge of the eyes, and its prevalence varies according to the patients’ gender and age, and different seasons of the year [16]. To the best of our knowledge, there is little evidence available in Iran about conjunctivitis caused by M. catarrhalis, although some studies have been performed on the bacterial and viral agents that cause the disease [17, 18]. We observed a higher occurrence of conjunctivitis in male (65%) when compared to female (35%) patients. A similar result was observed in an Italian independent study by Petrillo et al. [19]. In this study, the results of culture and PCR methods revealed the occurrence rate of 1% for M. catarrhalis in conjunctivitis samples. In a study by Afjeiee et al. [17] from Tehran, Iran, of the 241 neonates with clinical signs of conjunctivitis, the most frequent isolates were coagulase-negative Staphylococci (n = 130, 53.9%), followed by Chlamydia trachomatis (n =  40, 16.6%). They did not detect M. catarrhalis by routine culture method. Another report by Bhattacharyya et al. [20] from India revealed a frequency rate of 4.5% for M. catarrhalis among 110 culture- positive specimens of acute bacterial conjunctivitis that was higher than the current study. Also, Iwalokun et al. [21] from Nigeria reported an occurrence rate of 4.5% for Moraxella spp, among 83 conjunctival specimens.

So far, M. catarrhalis has been reported from several eye infection types, such as blepharitis, conjunctivitis, dacryocystitis, keratitis, and endophthalmitis [10]. The systematic review by Teweldemedhin et al. [10] documented a higher incidence of Moraxella spp., particularly M. catarrhalis in keratitis when compared to other eye infections. In a study by Jyoti et al. [22] from India, the prevalence of M. catarrhalis was reported as 0.4% in conjunctivitis specimens that was lower than this study. In previous reports from various countries of the world, the coagulase-negative Staphylococci and S. aureus were among the most prevalent species isolated from eye infections, including conjunctivitis [17, 19, 22].

Another point to consider about the bacterium M. catarrhalis is that its prevalence has rarely been evaluated by precise and sensitive molecular methods including PCR, real-time PCR, and loop-mediated isothermal amplification in ocular infections; while most studies of the presence of this bacterium in the respiratory specimens were done with molecular assessment in parallel with the conventional culture method [3, 23]. This may indicate that the actual prevalence of this bacterium in ocular infections is lower than the original rate, as the use of molecular methods is much more specific and sensitive than the conventional culture method. Some previous studies that used conventional culture method, matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), VITEK 2 system, and DNA sequencing have been reported several Moraxella species including M. nonliquefaciens, M. lacunata, M. osloensis, M. atlantae, and M. catarrhalis as a cause of ocular infections [3, 24, 25].

Conclusion

Although the M. catarrhalis was identified with a low rate as a causative agent of conjunctivitis in our region, this study can be considered as a starting point for investigation of this pathogen in all types of eye infections with larger sample size and with a multicentral design.

Limitation

In this study, the prevalence and incidence rates of other bacterial or viral pathogens that may have been responsible for conjunctivitis were not investigated due to the very low financial sources that can be considered as a limitation of the current research.

Availability of data and materials

The data of the current study are available from the corresponding authors on reasonable request.

Abbreviations

MALDI-TOF MS:

Matrix assisted laser desorption/ionization time-of-flight mass spectrometry

NTHi:

Nontypeable Haemophilus influenza

PCR:

Polymerase chain reaction

References

  1. 1.

    Shaikh SB, Ahmed Z, Arsalan SA, Shafiq S. Prevalence and resistance pattern of Moraxella catarrhalis in community-acquired lower respiratory tract infections. Infect Drug Resist. 2015;8:263–7.

    Article  Google Scholar 

  2. 2.

    Augustyniak D, Seredyński R, McClean S, Roszkowiak J, Roszniowski B, Smith DL, et al. Virulence factors of Moraxella catarrhalis outer membrane vesicles are major targets for cross-reactive antibodies and have adapted during evolution. Sci Rep. 2018;8(1):1–5.

    CAS  Article  Google Scholar 

  3. 3.

    LaCroce SJ, Wilson MN, Romanowski JE, Newman JD, Jhanji V, Shanks RM, et al. Moraxella nonliquefaciens and M. osloensis are important Moraxella species that cause ocular infections. Microorganisms. 2019;7(6):163.

    CAS  Article  Google Scholar 

  4. 4.

    Brown A, Addison B. Eye disorders: understanding the causes, symptoms and management. Pharm J. 2017;2:1–7.

    CAS  Google Scholar 

  5. 5.

    Yeu E, Hauswirth S. A review of the differential diagnosis of acute infectious conjunctivitis: implications for treatment and management. Clin Ophthalmol (Auckland, NZ). 2020;14:805–13.

    Article  Google Scholar 

  6. 6.

    Azari AA, Barney NP. Conjunctivitis a systematic review of diagnosis and treatment. JAMA. 2013;310(16):1721–9.

    CAS  Article  Google Scholar 

  7. 7.

    Sugita G, Hotomi M, Sugita R, Kono M, Togawa A, Yamauchi K, et al. Genetic characteristics of Haemophilus influenzae and Streptococcus pneumoniae isolated from children with conjunctivitis-otitis media syndrome. J Infect Chemo. 2014;20(8):493–7.

    CAS  Article  Google Scholar 

  8. 8.

    Raveendran S, Kumar G, Sivanandan RN, Dias M. Moraxella catarrhalis: a cause of concern with emerging resistance and presence of BRO beta-lactamase gene—report from a tertiary care hospital in south India. Int J Microbiol. 2020;2020:7316257.

    Article  Google Scholar 

  9. 9.

    Everitt HA, Little PS, Smith PWF. A randomized controlled trial of management strategies for acute infective conjunctivitis in general practice. BMJ. 2006;333:321–4.

    Article  Google Scholar 

  10. 10.

    Teweldemedhin M, Gebreyesus H, Atsbaha AH, Asgedom SW, Saravanan M. Bacterial profile of ocular infections: a systematic review. BMC Ophthalmol. 2017;17(1):212.

    Article  Google Scholar 

  11. 11.

    Tille PM. Bailey and Scott’s diagnostic microbiology. 13th ed. St. Louis: Elsevier; 2014.

    Google Scholar 

  12. 12.

    Yazdansetad S, Alkhudhairy MK, Najafpour R, Farajtabrizi E, Al-Mosawi RM, Saki M, et al. Preliminary survey of extended-spectrum β-lactamases (ESBLs) in nosocomial uropathogen Klebsiella pneumoniae in north-central Iran. Heliyon. 2019;5(9):e02349.

    Article  Google Scholar 

  13. 13.

    Montazeri EA, Khosravi AD, Saki M, Sirous M, Keikhaei B, Seyed-Mohammadi S. Prevalence of extended-spectrum beta-lactamase-producing Enterobacteriaceae causing bloodstream infections in cancer patients from southwest of Iran. Infect Drug Resist. 2020;13:1319–26.

    Article  Google Scholar 

  14. 14.

    Abbasi Montazeri E, Seyed-Mohammadi S, Asarehzadegan Dezfuli A, Khosravi AD, Dastoorpoor M, Roointan M, et al. Investigation of SCCmec types I–IV in clinical isolates of methicillin-resistant coagulase-negative staphylococci in Ahvaz, Southwest Iran. Biosci Rep. 2020;40(5):BSR20200847.

    Article  Google Scholar 

  15. 15.

    Post JC, Preston RA, Aul JJ, Larkins-Pettigrew M, Rydquist-White J, Anderson KW, et al. Molecular analysis of bacterial pathogens in otitis media with effusion. JAMA. 1995;273(20):1598–604.

    CAS  Article  Google Scholar 

  16. 16.

    Ryder EC, Benson S. Conjunctivitis. Treasure Island: StatPearls Publishing. 2020. https://www.ncbi.nlm.nih.gov/books/NBK541034/.

  17. 17.

    Afjeiee SA, Tabatabaei SR, Fallah F, Fard AT, Shiva F, Adabian S, et al. A microbiological study of neonatal conjunctivitis in two hospitals in Tehran, Iran. Asian Pac J Trop Dis. 2013;3(6):429–33.

    Article  Google Scholar 

  18. 18.

    Johari Moghadam MM, Yari MM, Jalilian FA, Amini R, Bazzazi N. Epidemiology and molecular diagnosis of acute conjunctivitis in patients attending Hamadan, west Iran ophthalmology clinics 2016–2017. Clin Optom. 2019;11:105–11.

    Article  Google Scholar 

  19. 19.

    Petrillo F, Folliero V, Santella B, Franci G, Foglia F, Trotta MC, et al. Prevalence and antibiotic resistance patterns of ocular bacterial strains isolated from pediatric patients in university hospital of Campania “Luigi Vanvitelli”, Naples, Italy. Int J Microbiol. 2020;2020:8847812.

    Article  Google Scholar 

  20. 20.

    Bhattacharyya A, Sarma P, Sarma B, Kumar S, Gogoi T, Kaur H. Bacteriological pattern and their correlation with complications in culture positive cases of acute bacterial conjunctivitis in a tertiary care hospital of upper Assam: a cross sectional study. Medicine. 2020;99(7):e18570.

    CAS  Article  Google Scholar 

  21. 21.

    Iwalokun BA, Oluwadun A, Akinsinde KA, Niemogha MT, Nwaokorie FO. Bacteriologic and plasmid analysis of etiologic agents of conjunctivitis in Lagos, Nigeria. J Ophthalmic Inflamm Infect. 2011;1(3):95–103.

    CAS  Article  Google Scholar 

  22. 22.

    Jyoti S, Kumar SA, Priyanka T, Nandan SB, Ramesh Y. Conjunctival microflora and their antibiotic susceptibility in north Indians prior to cataract surgery. Int J Curr Microbiol Appl Sci. 2014;3(9):254–9.

    Google Scholar 

  23. 23.

    Prates M, Tamashiro E, Proenca-Modena JL, Criado MF, Saturno TH, Oliveira AS, et al. The relationship between colonization by Moraxella catarrhalis and tonsillar hypertrophy. Can J Infect Dis Med Microbiol. 2018;2018:5406467.

    Article  Google Scholar 

  24. 24.

    Graham DR, Band JD, Thornsberry C, Hollis DG, Weaver RE. Infections caused by Moraxella, Moraxella urethalis, Moraxella-like groups M-5 and M-6, and Kingella kingae in the United Sates, 1953–1980. Rev Inf Dis. 1990;12:423–31.

    CAS  Article  Google Scholar 

  25. 25.

    Tobimatsu Y, Inada N, Shoji J, Yamagami S. Clinical characteristics of 17 patients with Moraxella keratitis. Sem Ophthalmol. 2018;33:1–7.

    Article  Google Scholar 

Download references

Acknowledgements

We are thanking all personal and resident students of the Ophthalmology Division and laboratory ward of Emam Khomeini Hospital.

Funding

This study was financially supported by the Research Affairs of Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran.

Author information

Affiliations

Authors

Contributions

AFS and MF conceived the project and designed the experiments. MF, MT, MS, and HV designed and collected samples. MT, MS, and HV analyzed the data. AFS and MF supervised the collection of the samples. AFS wrote the manuscript. All authors reviewed the manuscript. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Morteza Saki or Hojat Veisi.

Ethics declarations

Ethics approval and consent to participate

This research was approved by the Microbiology Department of Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran based on the Declaration of Helsinki. Written informed consent was obtained from all patients.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Farajzadeh Sheikh, A., Feghhi, M., Torabipour, M. et al. Low prevalence of Moraxella catarrhalis in the patients who suffered from conjunctivitis in the southwest of Iran. BMC Res Notes 13, 547 (2020). https://doi.org/10.1186/s13104-020-05389-4

Download citation

Keywords

  • Moraxella catarrhalis
  • Conjunctivitis
  • Ocular infection
  • Iran