Mycoplasma pneumoniae is a free living micro-organism which is classified in the family Mycoplasmataceae. It is primarily a human respiratory pathogen and the infection may vary from a mild pharyngitis to a pneumonia. M. pneumoniae infections are usually endemic in the community but periodic epidemics can also occur. These micro-organisms are sensitive to specific antibiotics such as tetracycline and erythromycin and a laboratory diagnosis using rapid methods is therefore important. Studies were carried out at the Virology Laboratory, St. Luke’s Hospital involving 1,022 cases during the period 1995 to 1998. These were examined for specific anti-mycoplasmal IgM antibodies using an enzyme immunoassay (EIA) technique. One hundred and forty samples or 13.7% were found to be reactive. Compared to the traditional cultural methods, EIA techniques are very rapid laboratory diagnostic methods, the result being communicated to the clinician within a few hours. The differential diagnosis of the particular case can thus be resolved quickly and appropriate antibiotic therapy instituted without much delay for the maximum benefit to the patient.peer-reviewe

Barbara, Christopher

Portelli, Alfred V.

English

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Maltese Medical Journal, 2000; 12(1,2): 40 All rights reserved Mycoplasma Pneumoniae infections in the Maltese islands A. Portelli*, C. Barbara* * This article is based on a paper presented at the IVth Maltese Medical School Conference held at the Mediterranean Conference Centre on 10_14th March 1999. ABSTRACT: Mycoplasma pneumoniae is a free living micro-organism which is classified in the family Mycoplasmataceae. It is primarily a human respiratory pathogen and the infection may vary from a mild pharyngitis to a pneumonia. M. pneumoniae infections are usually endemic in the community but periodic epidemics can also occur. These micro-organisms are sensitive to specific antibiotics such as tetracycline and erythromycin and a laboratory diagnosis using rapid methods is therefore important. Studies were carried out at the Virology Laboratory, St. Luke's Hospital involving 1,022 cases during the period 1995 to 1998. These were examined for specific anti-mycoplasmal IgM antibodies using an enzyme immunoassay (EIA) technique. One hundred and forty samples or 13.7% were found to be reactive. Compared to the traditional cultural methods, EIA techniques are very rapid laboratory diagnostic methods, the result being commu­nicated to the clinician within a few hours. The differential diagnosis of the particular case can thus be resolved quickly and appropriate antibiotic therapy instituted without much delay for the maximum benefit to the patient. 'Department of Pathology, University of Malta Medical School, Gwardamangia, Malta "Department of Pathology, Virology Laboratory, St. Luke's Hospital, Gwardamangia, Malta Correspondence: Dr. Alfred Portelli, Department of Pathology, University of Malta Medical School, Gwardaman­gia, Malta. E-mail: alfport@hotmail.com Keywords: Mycoplasma pneumoniae, rapid laboratory methods, specific antibiotics Introduction encephalitis7. M. pneumoniae forms pleomorphic filaments with an Mycoplasma pneumoniae, also called Eaton's agent average diameter of about 0.51Am and lacks the pepti­after the investigator who discovered it, is a free-living doglycan cell wall component characterisitic of bacteria. micro-organism which is classified in the genus My­ The absence of the peptidoglycan makes this micro­coplasma in the family Mycoplasmataceae '. It is a hu­ organism resistant to penicillins, cephalosporins and man pathogen and mainly causes respiratory diseases other antibiotics which interfere with the integrity of ranging from mild pharnygitis to tracheobronchitis and this structure. The cytoplasmic contents are enclosed in pneumonia2• The latter infection is usually insidious in a well-developed plasma membrane. The major anti­onset with generalised malaise, pyrexia and headache. genic determinants are present in this membrane and are The symptoms usually increase within a couple of days composed of glycolipids and proteins. Cross-reactivity and a hacking non-productive cough develops. Tired­ of these antigens with human tissues and other micro­ness and marked weakness are also common. A maculo­ organisms occurs. M. pneumoniae is fastidious and papular rash may appear in about 15% of cases. Muscle grows slowly on complex enriched media giving rise to tenderness and arthralgias are common in all age colonies which have a homogenous granular morphol­groups. The infection is seen especially in older children ogy unlike the "fried-egg" appearance of other my-and young adults, being milder and asympto­Jan-Mar Apr-Jun Jul-Sep Oct-Decmatic in pre-school children3.4. M. pneumo­niae infections are endemic in the community Reactive 38 33 19 27 but epidemics occur every 4-5 years. Non­ Samplesrespiratory manifestations caused by M. Non Reactive 348 243 157 163pneumoniae include mucocutaneous Sampleseruptions and haemolytic anaemias the latter being possibly due to the development of Total No. of 386 276 176 190 'cold agglutinins ', pericarditis and myocardi­ samples tested tis6 and neurological involvement such as Table 1 - Total Mycoplasma samples according to season meningism, aseptic meningitis or meningo­42 A. Portelli, C. Barbara unknown origin. The patients' age varied from infants of a few months to elderly people in their eighties. The test chosen for the laboratory diagnosis was an01995 enzyme immunoassay test for the detec­01996 tion of anti-mycoplasma IgM antibodies .1997 and adapted to microtitre plate technology Ill] 1998 employing the reverse EIA technique (Savyon Diagnostics Ltd, Israel. 1994). Results -	Jan-Mar Apr-Jun Jul-Sep Oct-Dec Trimesters 	 Out of 1,022 samples examined, 140 or 13.7% were found to be reactive to M. pneumoniae. Table 1 and Figures depict Fig 1 - Reactive Mycoplasma samples according to season (n=117) the reactive samples according to season, age group, sex and locality. The data in coplasmas. The colonies are very small and details can each Figure may not necessarily add to the total number only be seen with a colonial microscope. Unlike other of reactive samples because the data on some cards sub­species which are facultatively anaerobic, M. pneumo­ mitted to the laboratory with the samples was missing or niae grows aerobicalll. the writing was illegible. Table 1 and Figure 1 show that Although mycoplasmas are considered to be bacteria, out of 117 reactive samples, 38 were positive during the the laboratory diagnosis of this 'paraviral' is traditionally January-March period, 33 in the second period, 19 in carried out in a virology laboratory. Besides cultural the third and 27 during the October-December period. methods, the serological assays most frequently em­ Figure 2 shows 99 reactive samples according to age ployed are the cold agglutination and complement fixa­ groups. Twenty-five were in the 0 to 5 years age group, tion tests and enzyme immunoassays. 41 in the 6 to 20 years age group while 33 were present in the group 21 years and over. Table 2 and Figure 3 Materials and Methods show the number of reactive samples according to sex. Fifty-six out of 114 samples were from males while 58 Studies were carried out at the Virology laboratory, St. were from females. Figure 4 shows locality distribution Luke's Hospital. All requests for the detection of My­ with three zones, North, Central and South, dividing the coplasma IgM antibodies during the four-year period Maltese population into approximately three equal 1995 to 1998 were entered into a database. Three per­ population zones. Twenty-six samples came from pa­cent of the samples had to be excluded because they tients living in the North part of the islands including lacked essential data or the samples were haemolysed. Gozo. Thirty-four samples came from the Central region Repeats were also excluded. One thousand and twenty­ while 33 reactive samples originated from the South. two samples were included and of these 52% were from As in similar studies9 local M. pneumoniae infections male and the rest from female patients. Ninety-four per­ were found to occur throughout the year with no consis­cent of all referrals were from general hospitals (St. tent increased seasonal variation, although in our study, Luke's and Gozo General Hospital), 4% were from out­ there was a slight increase in the number of reactive patients ' clinics and only 2% were referred from the cases in the winter of 1997 (Fig. 1). Disease was more community (health centres and general practice). The common in the younger age group (Fig. 2) and there majority of samples were obtained from patients suffer­ were no overall differences between the sexes (Fig. 3) ing from respiratory disease or from cases of pyrexia of or between the different localities of the Maltese Is­lands (Fig. 4). Discussion Age Groups Culture for M. pneumoniae can be carried OOto 5 out for the laboratory diagnosis of M. D6 to 20 pneumoniae infection but growth is slow .21+ and the result can only be expected in about 2 to 6 weeks lO • The cold agglutina­tion test is carried out by reacting the pa­Z tient's serum with human group 0 erythro­1995 1996 1997 1998 cytes incubated at 4 0c. It appears that M. pneumoniae possesses a surface antigenYears which is similar to the I antigen of these red blood cell types. A positive cold agglu­Fig 2 - Reactive Mycoplasma samples according to age group (n=99) tination assay is observed, on average, in le 20 0.. ~ 15 In Cl) ~ 10 ca ~ 5 '0 o 0z le 15 0.. E ca In 10 Cl) >;:: u 5ca Cl)... '0 0 0 43 '. , Mycoplasma Pneumoniae infections in the Maltese islands Fig 3 - Reactive Mycoplasma samples according to sex (n=114) Acknowledgements We would like to thank the non-medical staff of the Virology Laboratory at St. ~ 16~------------------------~ Fig 4 - Reactive Mycoplasma samples according to locality (n=93) infections in children. Arch Dis Child. 53: 38­Non reactive samples Reactive samples Total I samples tested Males Females Table 2 - Total Mycoplasma samples according to sex ~ 25 ;>-~-~-~.~~.-"-.~-~.""--~~""",...~,,,,., 15.. E 20 -I--------i ca III Cl) 15 -1--------; > ~ 10ca Cl) ... 5 '0 o z o 1995 1996 1997 1998 Years 4. only 50% of patients with M. pneumoniae infection. The test is not specific for M. pneumonaie and cross­5.reactions occur with other infective agents e.g. Epstein­Barr virus and Adenovirusll . The complement fixation test utilises a lipid extract of 6. the pathogen but this has been shown to cross-react with components of human tissue as well as bacterial glycol­ipids. It requires complicated reagent titrations which 7. makes this test somewhat difficult to perform even by experiencep laboratory workers. The test is usually sig­8. 1995 1996 1997 1998 188 365 224 132 22 37 28 27 210 402 252 159 105 210 114 81 95 192 138 78 I-~~ : 10 > g 6 e 4 '0 2 o oz 1995 1996 1997 Years n• 8 +----1 '1-----1 "." Luke's Hospital for their technical assis­tance and without whose help this study would not have been possible. El North El Central References • South 1. Tully IG. Current status of the mollicute flora off humans. Clin Infect Dis 17(Suppl. 1): 2-9, 1993. 2. Clyde W A. Clinical overview of typical1998 Mycoplasma pneumoniae infections. Clin Infect Dis 17 (Suppl. 1): 32-36, 1993. 3. Stevens D, Swift PFG, Keamey PI, Cor­ner BD, Burman D. Mycoplama pneumoniae nificant if rising titres of antibodies can be demonstrated in two consecutive specimens taken at least 10 days apart and, therefore, introducing a delay in diagnosis. The EIA test employed in our study is a rapid method of diagnosis, the result being transmitted to the remit­ting medical officer in a few hours. The test is also eas­ier to perform than the complement fixation test and it shows higher sensitivity and specificity12. Although this study included all the referrals for My­coplasma antibody tests, the total number of samples was small and, correspondingly, the number of positives was also small. It is therefore difficult to draw firm con­clusions and perhaps a larger follow-up study is indi­cated. Conclusion A rapid method has been utilised to carry out the laboratory diagnosis of M. pneu­moniae infections. Since this micro­organism is usually sensitive to antibiot­El Male ics of the tetracycline and macrolide • Female classes, it follows that appropriate ther­apy can be started without much delay and therefore to the maximum benefit of the patient. It also follows that clinicians should keep Mycoplasma pneumoniae in mind whenever the differential diagnosis of a respiratory infection is considered. 42.1978. Murray HW, Masur H, Senterfit LB, Roberts RB. The protean manifestations of M. pneumoniae infections in children. Clin Infect Dis. 17(Suppl1.): 47-51, 1993. Cherry ID. Anaemia and mucocutaneous lesions due to Mycoplasma pneumoniae infections. Clin Infect Dis 17 (Suppl1.): 47-51, 1993. Kenny RT, Li IS, Clyde WA et al. Mycoplasma pericar­ditis: evidence of invasive disease. Clin Infect Dis 17 (Suppl. 1): 58-62,1993. Narita M, Matsuzono Y, Togashi T et al. DNA diagnosis of central nervous system infection by Mycoplasma pneumoniae. Paediatrics 90: 250-253, 1992. Razin S, Barile MF. The Mycoplasmas, vol. 4. Aca­45 Mycoplasma Pneumoniae infections in the Maltese islands demic Press, Inc., New York. 1985. 9. 	 Foy HM, Kenny GE, Cooney MK and Alien ID. Long term epidemiology of infections with M. pneumonaie. J Infect Dis 139: 681-687,1979. 10. 	 Waites K et al. Laboratory diagnosis of mycoplasmal and urea-Iasmal infections. Clin Microbiol. Newslett. 18: 105-112, 1996. 11. 	 Jacobs E. Serological diagnosis of Mycoplasma pneu­---. moniae infections: a critical review of current proce­dures. Clin Infect Dis 17(Suppl. 1): 79-82, 1993. 12. 	 Dussaix E, Slim A, Toumier P. Comparison of en­zyme-linked immunosorbent assay (ELISA) and com­plement fixation test for detection of Mycoplasma pneumoniae antibodies. J Clin Pathol. 36: 228-232, 1983. The copyright of this article belongs to the Editorial Board of the Malta Medical Journal. The Malta Medical Journal’s rights in respect of this work are as defined by the Copyright Act (Chapter 415) of the Laws of Malta or as modified by any successive legislation. Users may access this full-text article and can make use of the information contained in accordance with the Copyright Act provided that the author must be properly acknowledged.  Further distribution or reproduction in any format is prohibited without the prior permission of the copyright holder. This article has been reproduced with the authorization of the editor of the Malta Medical Journal (Ref. No 000001)  

Anaemia and mucocutaneous lesions due to Mycoplasma pneumoniae infections.

Comparison of en zyme-linked immunosorbent assay (ELISA) and com plement fixation test for detection of Mycoplasma pneumoniae antibodies.

Cooney MK and Alien ID. Long term epidemiology of infections with

DNA diagnosis of central nervous system infection by Mycoplasma pneumoniae.

Laboratory diagnosis of mycoplasmal and urea-Iasmal infections.

Mycoplasma pericar ditis: evidence of invasive disease.

Serological diagnosis of Mycoplasma pneu ---. moniae infections: a critical review of current proce dures.

The protean manifestations of M. pneumoniae infections in children. Clin Infect Dis.

Mycoplasma Pneumoniae infections in the Maltese Islands

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Mycoplasma Pneumoniae infections in the Maltese Islands

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