Evaluation of colonization, variable lipoprotein-based serological response, and cellular immune response of Mycoplasma hyorhinis in experimentally infected swine.

Mycoplasma hyorhinis (Mhr) is a commensal of the upper respiratory tract that can be shed by nasal secretions and transmitted by direct contact in neonatal and nursery pigs. Lesions associated with Mhr infection include polyserositis and arthritis; however, systemic Mhr disease pathogenesis is not well characterized. This study aimed to investigate the immunopathogenesis and bacterial dissemination pattern of Mhr using single and multiple inoculation approaches in a caesarian-derived colostrum-deprived (CDCD) pig model. Animals in three treatment groups were inoculated once (Mhr 1; n = 12) or four (Mhr 2; n = 8) times with Mhr or sham-inoculated (NC group; n = 3) nasally and by tonsillar painting. Inoculum consisted of a triple cloned Mhr field isolate (4.5 × 107 CFU/mL) in Friis medium. Clinical signs were evaluated daily during the study. Serum and oral fluid antibody (IgA and IgG) response and cellular immune response were assessed using a recombinant chimeric VlpA-G-based indirect ELISA and by ELISpot, respectively. The presence of Mhr in oral fluids, nasal and oropharyngeal swabs were evaluated by qPCR. At 6 wpi, pigs were euthanized and evaluated for gross lesions consistent with Mhr and bacterial colonization in tonsils by qPCR. No clinical signs or gross lesions consistent with Mhr-associated disease were observed throughout the study. For Mhr 2 group, the presence of IgA and IgG in serum and oral fluids were detected at 2 and 4 weeks post-inoculation (wpi), respectively, while in Mhr 1, only IgA was detected in oral fluids at 6 wpi. The proportion of animals shedding Mhr in nasal secretions varied from 20 to 40 % in the Mhr 1 and 62.5-100% in the Mhr 2 group. However, the proportion of animals shedding Mhr in oropharyngeal swabs was consistent through the study (60 %) in Mhr 1 and fluctuated from 20 % to 87.5 % in Mhr 2 group. The lack of clinical signs and the presence of Mhr specific humoral response and bacterial colonization indicates that the multiple inoculation experimental model may mimic subclinical natural infection in the field. In addition, the humoral and transient cellular response did not result in bacterial clearance. Based on these results, animals would have to be exposed multiple times to mount a detectable immune response.

Molecular detection of Mycoplasma synoviae and avian reovirus infection in arthritis and tenosynovitis lesions of broiler and breeder chickens in Santa Catarina State, Brazil.

Infectious arthritis or tenosynovitis in broiler and breeder chickens results in major loss of productivity because of reduced growth and downgrading at processing plants. The most common causative agents of avian infectious arthritis are the bacterium Mycoplasma synoviae and avian reoviruses (ARVs) (family Reoviridae, genus Orthoreovirus). In this study, we evaluated the occurrence of these two pathogens in arthritis or tenosynovitis lesions of broilers and breeder flocks in southern Brazil using molecular detection. Tissue sections from tibiotarsal joints with visible lesions from 719 broilers and 505 breeders were analysed using pathogen-specific polymerase chain reaction (PCR) assays. In breeders, 41.2% (n = 296) of lesions were positive for M. synoviae, 26.4% (n = 190) were positive for ARV, while co-infection was present in 12.2% (n = 88) of the samples. In broilers, 20.8% (n = 105) of lesions were positive for M. synoviae, 11.9% (n = 60) for ARV and 7.7% (n = 39) of these cases were positive for both pathogens. Post-mortem examination revealed lesions with varying degrees of gross pathological severity. Histopathological examination showed intense, diffuse lymphohistiocytic inflammatory infiltrates with heterophil accumulation, primarily in the synovial capsule and digital flexor tendon, in all samples. Improved strategies for early detection and control of these major avian pathogens are highly desirable for preventing the spread of infection and reducing economic losses in the poultry industry.

Suppression of Baeckea frutescens L. and its components on MyD88-dependent NF-κB pathway in MALP-2-stimulated RAW264.7 cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Baeckea frutescens L. is commonly used as a folk medicinal material. There are nineteen components in its volatile oil, including Pcymol which has effects of eliminating phlegm, relieving asthma and antiviral. This study was aimed to investigate the anti-infectious inflammatory activities of Baeckea frutescens L. and its conponents and analyzing the mechanisms. MATERIALS AND METHODS: The anti-infectious inflammation of Baeckea frutescens L. were studied by using macrophage activating lipopeptide-2 (MALP-2)-stimulated RAW264.7 cell model in vitro. Secretion of nitric oxide (NO), expression of inducible NO synthase (iNOS) and cytokines were detected as classic inflammatory index. Expression of Myeloid differentiation factor 88 (MyD88), degradation of inhibitory κBα (IκBα) and nuclear translocation of NF-κB p65 were further investigated. RESULTS: The results suggested that Baeckea frutescens L. has effect on suppression of MALP-2-mediated inflammation in RAW264.7 cells. The secretion of NO and the expression of iNOS could be inhibited. The secretion of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) were also declined. Baeckea frutescens L. significantly decreased the expression of MyD88, therefore, inhibited the degradation of IκBα, reduced the level of nuclear translocation of p65. CONCLUSION: The results of this study indicated that Baeckea frutescens L. and its components could inhibit the anti-infectious inflammatory events and iNOS expression in MALP-2 stimulated RAW264.7 cells. Among them, BF-2 might play a role through the inhibition of the MyD88 and NF-κB pathway. Our study might provide a new strategy to design and develop this kind of drug towards mycoplasma-infected inflammation.

Patho-bacteriological investigation of an outbreak of Mycoplasma bovis infection in calves - Emerging stealth assault.

Mycoplasma bovis (M. bovis) is an important bacterium, causing severe respiratory infection, and arthritis in dairy animals worldwide. This study is based on 50 suckling calves among which 15 showed respiratory distress, lameness and swollen joints and died later. M. bovis was isolated and identified from all dead (n = 15) and live (17.14%; 06 out of 35) calves on the basis of bacteriological examination. In morbid calves, the carpus and stifle joints were severely affected, while necropsy revealed multiple well-circumscribed calcified abscesses and caseous exudates in cranio-ventral and diaphragmatic lobes of lungs. Suppurative polyarthritis, fibrino-suppurative, teno-synovitis and otitis media were the common and striking lesion observed at postmortem examination. Histopathological examination revealed broncho-interstitial pneumonia and necrotic fibrino-purulent broncho-pneumonia in lungs. Similarly, synovial membranes and joints revealed presence of multiple foci of liquefactive necrosis surrounded by lymphocytes, plasma cells, macrophages and peripheral fibroplasia. In the bacteriological investigations, the characteristic fried egg colonies of M. bovis further confirmed this infection in all suspected cases. In conclusion, the current clinico-histo-pathological features are the depictive picture, and is the first report of M. bovis infection in calves in Pakistan.

Mycoplasma corogypsi-associated polyarthritis and tenosynovitis in black vultures (Coragyps atratus).

Three wild American black vultures (Coragyps atratus) were presented to rehabilitation centers with swelling of multiple joints, including elbows, stifles, hocks, and carpal joints, and of the gastrocnemius tendons. Cytological examination of the joint fluid exudate indicated heterophilic arthritis. Radiographic examination in 2 vultures demonstrated periarticular soft tissue swelling in both birds and irregular articular surfaces with subchondral bone erosion in both elbows in 1 bird. Prolonged antibiotic therapy administered in 2 birds did not improve the clinical signs. Necropsy and histological examination demonstrated a chronic lymphoplasmacytic arthritis involving multiple joints and gastrocnemius tenosynovitis. Articular lesions varied in severity and ranged from moderate synovitis and cartilage erosion and fibrillation to severe synovitis, diffuse cartilage ulceration, subchondral bone loss and/or sclerosis, pannus, synovial cysts, and epiphyseal osteomyelitis. No walled bacteria were observed or isolated from the joints. However, mycoplasmas polymerase chain reactions were positive in at least 1 affected joint from each bird. Mycoplasmas were isolated from joints of 1 vulture that did not receive antibiotic therapy. Sequencing of 16S rRNA gene amplicons from joint samples and the mycoplasma isolate identified Mycoplasma corogypsi in 2 vultures and was suggestive in the third vulture. Mycoplasma corogypsi identification was confirmed by sequencing the 16S-23S intergenic spacer region of mycoplasma isolates. This report provides further evidence that M. corogypsi is a likely cause of arthritis and tenosynovitis in American black vultures. Cases of arthritis and tenosynovitis in New World vultures should be investigated for presence of Mycoplasma spp, especially M. corogypsi.

Recent perspectives in the diagnosis and evidence-based treatment of Mycoplasma genitalium.

Mycoplasma genitalium is a globally important sexually transmitted pathogen. Men infected with M. genitalium frequently present with dysuria, while women may present with or without urogenital symptoms. In some populations, M. genitalium is significantly associated with HIV-1 infection, and is also an etiological agent in pelvic inflammatory disease. However, there is insufficient evidence to establish a causative role of the organism in obstetric complications, including tubal factor infertility. Although several nucleic acid amplification tests offer rapid, sensitive methods for detecting M. genitalium, there is no standardized assay. Available evidence supports treatment of M. genitalium infections with an extended regimen of azithromycin and resistant strains respond to moxifloxacin. Accumulating evidence indicates growing fluoroquinolone resistance, including against moxifloxacin, emphasizing the need for new therapeutic strategies to treat M. genitalium infections.

Successful treatment of multiple subdural empyemata caused by Mycoplasma hominis in a newborn.

We report a case of neonatal subdural empyema caused by Mycoplasma hominis. The infant sustained severe birth-related eye injury and subsequently developed multiple subdural empyemata. This report illustrates the clinical presentation of this unusual infection and the crucial role of neurosurgical intervention and specific antimicrobial therapy for its successful management.

Mycoplasma pneumoniae-associated mucositis with minimal skin manifestations.

Mycoplasma pneumoniae-associated mucositis is a rarely described complication of M. pneumoniae infection presenting with ocular, oral, and genital involvement but without the typical skin lesions seen in Stevens-Johnson syndrome. A 27-year-old man with a past history of asthma presented at the emergency room with a 1-week history of cough (initially non-productive but subsequently associated with non-bloody mucopurulent sputum), fever, myalgias, headache, and progressive dyspnea. Two days before admission he had commenced amoxicillin/clavulanic acid with no improvement. The patient reported bilateral conjunctival injection and hemorrhagic ulcers on the lips commencing the day prior to admission. Physical examination revealed fever (39 degrees C), bilateral exudative conjunctivitis, painful hemorrhagic ulcers on the lips, tongue, and oral mucosa, small scrotal erosions, erythema of the penile meatus, and small erythematous bullae on the dorsum of each hand; subsequently, the patient developed bullae at the venipuncture site on his right arm. Laboratory tests revealed positive IgM serology for M. pneumoniae, with titer elevation. The patient was successfully treated with levofloxacin and prednisolone. Our case appears to be the first adult patient described with M. pneumoniae-associated mucositis, which has previously been reported only in pediatric patients. This is also the first reported instance of a case of M. pneumoniae-associated mucositis treated with levofloxacin and prednisolone. M. pneumoniae infection should be considered in all cases of mucositis, and treatment of this condition with levofloxacin and prednisolone seems to be effective.

Suppurative polyarthritis in striped skunks (Mephitis mephitis) from Cape Cod, Massachusetts: detection of mycoplasma DNA.

Striped skunks (Mephitis mephitis) from Cape Cod, Massachusetts, U.S.A. were necropsied (n=34; 1995-1997) or clinically evaluated (n=25, 2002-2003) to characterize a lameness and polyarthritis, reported by wildlife veterinarians and rehabilitators, and unsuccessfully treated with antibiotics. Overall, 22 affected skunks had one or multiple swollen joints, swollen paws, and subcutaneous abscesses. Purulent exudate was located in joint spaces, in periarticular connective tissue between muscle fascicles and tendons, and between and along flexor and extensor tendons of the paws. Histologic examination revealed suppurative arthritis, with necrosis and erosion of articular cartilage, and suppurative osteomyelitis. Special stains failed to reveal a causative microorganism within affected joints, and routine bacteriologic cultures failed to isolate a pathogen with any significant frequency or consistency. Polymerase chain reaction (PCR) experiments were performed using DNA extracted from archived, formalin-fixed joint samples of 11 affected skunks, and DNA from joints of 7 of 11 affected skunks yielded amplicons with sequences highly similar to sequences of Mycoplasma fermentans within the Mycoplasma bovis cluster, whereas DNA samples from joints of four unaffected skunks were negative by PCR. Skunks from Connecticut, U.S.A. (n=21; 1995-2003) were similarly examined and were found not to have suppurative polyarthritis, suggesting a unique geographic distribution of this condition. Concurrent pathologic conditions in adult skunks from both Cape Cod and Connecticut included verminous pneumonia, gastric nematodiasis, arthropod ectoparasitism, and canine distemper. Amyloidosis was present in skunks with and without suppurative polyarthritis, and the amyloid was immunohistochemically identified as AA-amyloid. This is the first report of suppurative polyarthritis in wild skunks with evidence of a mycoplasmal etiology.

The efficacy of valnemulin (Econor) in the control of disease caused by experimental infection of calves with Mycoplasma bovis.

Mycoplasma bovis infection was experimentally induced in groups of six young calves. A further group was uninfected and served as a control. Ten days after infection, medication with either enrofloxacin (Baytril, Bayer) or valnemulin (Econor, Novartis) was instituted via the milk replacer for a further 10 days, after which all calves were killed. Infection resulted in depression, pyrexia, inappetance and prominent respiratory signs. Arthritis occurred in two animals and two (unmedicated) animals died. At post-mortem examination extensive lesions were present in the lungs and M. bovis was re-isolated from infected unmedicated calves' lungs. Medication with either enrofloxacin or valnemulin resulted in a rapid diminution of clinical signs, restoration of appetite and reversal of weight loss. Isolation of Pasteurella multocida from the calves' lungs was suppressed by both medicaments. Valnemulin resulted in a more rapid reduction of clinical scores and eliminated M. bovis from the lungs more effectively than enrofloxacin.

Testing the efficacy of fermented wheat germ extract against Mycoplasma gallisepticum infection of chickens.

The effect of fermented wheat germ extract (FWGE, Immunovet-HBM) was studied in chickens challenged with Mycoplasma gallisepticum. Ninety M. gallisepticum- and M. synoviae-free 3-wk-old chickens were exposed to aerosol infection of M. gallisepticum. One group (30 birds) was treated with FWGE, a second group with tiamulin, and a third group was untreated. The fourth group was exposed to PBS aerosol as a negative control. On d 9, all chickens were slaughtered and examined for the presence of gross and histological lesions, the presence of the challenge strain in the organs and specific antibodies in the serum. Body weight gains and feed conversion rates were recorded. In the groups treated with FWGE and with tiamulin, the chickens remained clinically healthy: their BW gains were 441.7 g and 446.8 g, respectively. Feed conversion ratios were 1.72 and 1.71 for FWGE- and tiamulin-treated birds, respectively. Control birds had BW gain of 480.8 g, and feed conversion ratio of 1.78. The numbers of birds with gross lesions (15 and 11, respectively) and lesion scores (25 and 25, respectively) of the FWGE- and tiamulin-treated groups were significantly lower than in the infected untreated group (25 birds, lesion score of 190). No mycoplasma was reisolated from brain, liver, spleen, heart, or kidneys of the FWGE-treated birds, and the number of mycoplasma isolations from the respiratory tract samples was less frequent (10) than from the infected untreated group (64). In addition, 35 samples from other internal organs were also positive. Twenty percent of the birds treated with FWGE showed serological response with a 5.0% reaction score, whereas in the infected untreated group, 83.3% of birds were reactors, with a 62.5% reaction score.

[Bronchopneumonia and polyarthritis due to Mycoplasma bovis in a calf]. / Bronchopneumonie und Polyarthritis mit Nachweis von Mycoplasma bovis bei einem Rind.

This case report describes gross lesions and histopathological findings in a 3-months-old calf originating from a feedlot with approximately 400 cattle. In this animal and additional 14 cattle of similar age, which were kept together in the same stable, swollen joints had occurred suddenly. The examination of this calf showed that a severe polyarthritis induced by haematogenous spread of Mycoplasma bovis following bronchopenumonia was present, which was characterised by necrotising lesions of the joint capsules and severe cartilage erosions.

Sensory properties of articular afferents following Mycoplasma arthritis in the chicken.

The physiological properties of joint capsule mechanical nociceptors of monoarthritic chickens (Gallus domesticus) were studied by recording the electrical activity from single C (Group IV) and A-delta (Group III) fibres dissected from the parafibular nerve. By injecting live Mycoplasma gallisepticum cultures into the ankle joint a typical mycoplasma arthritis was induced which was restricted to a single joint. During the early stage of the disease (7-21 days after infection) there was histopathological evidence of an acute synovitis and the fibres showed evidence of sensitisation. Sensitisation was observed in the significantly increased receptive field size, decreased response thresholds, increased response to joint movement both noxious and innocuous, but only in the C-fibres was there an increase in spontaneous activity. During the more chronic stage of the disease (49-56 days after infection) there was pathological evidence of prolonged synovitis but the sensory fibres responded normally to mechanical stimulation and joint movement. These changes in sensitivity of the joint capsule mechanical nociceptors provides peripheral neural evidence of possible pain experienced during the acute stage of the disease but not at the chronic stage when the disease might be in period of remission. The absence of any clear correlation between pathology and receptor activity demonstrates the difficulty of trying to predict nociceptive consequences in animals on the basis of histopathology.

Adherence to various host cell lines of Mycoplasma bovis strains differing in pathogenic and cultural features.

Mycoplasma bovis is known to be responsible for pneumonia and arthritis in calves, as well as mastitis in dairy cows. Despite clear evidence of its pathogenic potential, little is known about mechanisms of cytadherence and the molecular factors involved. The purpose of this work was to compare adherence rates of M. bovis field strains to different host cell lines and study the effects of cloning and sub-culturing M. bovis strains on their adherence properties. Eighteen metabolically labeled M. bovis strains isolated from different pathological backgrounds were examined in adherence trials using four different host cell lines, i.e. embryonic bovine lung (EBL), embryonic bovine trachea (EBTr), Madin Darby bovine kidney (MDBK) and rabbit kidney (RK) cells. Although large interstrain variations in adherence rates (3.4-19.1%) were measured they could not be correlated to the pathological background (pneumonia, arthritis or mastitis). Adherence rates to the fibroblast cell line (EBTr) were significantly lower than those to the three epithelial cell lines (EBL, MDBK and RK). The only non-pathogenic strain (221/89) exhibited lower adherence rates than three isolates from clinical mastitis. Interestingly, adherence rates were significantly reduced after in vitro passaging. In contrast, no effect of single cloning of strains on adherence was observed. There was no general correlation between expression of variable surface proteins (Vsps) as monitored by immunoblotting and adherence rates, although alterations in Vsp expression profiles were seen as a consequence of passaging. As there is probably a large number of adhesins, variable and non-variable, on the surface of M. bovis cells the issue is very complex, and the most active components have yet to be identified.

Role of bacteriophage MAV1 as a mycoplasmal virulence factor for the development of arthritis in mice and rats.

The lysogenic bacteriophage MAV1 has been shown to be a virulence factor for the development of arthritis in rats infected with Mycoplasma arthritidis. In the present study, arthritis was evaluated by histopathologic examination to demonstrate that MAV1 is a virulence factor not only in the rat but also in the mouse. Specifically, the MAV1 lysogen 158L3-1 was more virulent than the nonlysogen strain 158 in DBA/2NCr, C3H/HeNCr, C3H/HeJ, and C3Smn.CB17-Prkdc(scid)/J mice, as well as in LEW rats.

Sensory denervation by neonatal capsaicin treatment exacerbates Mycoplasma pulmonis infection in rat airways.

Mycoplasma pulmonis infection in rats results in life-long disease, characterized by chronic inflammation of the airway mucosa with widespread accumulation of lymphoid tissue, mucous cell hyperplasia, and mucosal thickening. In addition, there is angiogenesis and increased sensitivity of mucosal blood vessels to substance P (SP), so tachykinins released from sensory nerve fibers cause an abnormally large amount of plasma leakage. We sought to learn whether the sensory nerves influence the severity of the chronic inflammatory response of M. pulmonis infection. Our strategy was to destroy the nerves by capsaicin pretreatment at birth, infect the rats with M. pulmonis at 8 wk of age, and then study the animals 6 wk later. We found that capsaicin pretreatment increased the severity of the infection, exaggerated the pathological changes in the tracheal mucosa, and increased the amount of SP-induced plasma leakage, as quantified with Monastral blue. The thickness of the tracheal mucosa in these infected rats was 80% greater than in their vehicle-pretreated counterparts and 200% greater than in the pathogen-free controls. The area density of Monastral blue-labeled blood vessels averaged 20% in the infected rats pretreated with capsaicin, which represented a 40-fold increase over the leakage in the pathogen-free group. By comparison, the amount of Monastral blue labeling was only 13% in rats pretreated with vehicle (P<0.05), which was a 22-fold increase over the corresponding pathogen-free group. The number of SP-immunoreactive nerve fibers was reduced both by neonatal capsaicin and by infection (87 and 63% reductions, respectively); but when the two conditions were combined, their effects were not additive (79% reduction), perhaps because of nerve regrowth. We conclude that destruction of sensory nerves increases the severity of infection- induced chronic inflammation in the airway mucosa, with exaggerated mucosal thickening, angiogenesis, plasma leakage, and nerve remodeling.

Ciliostasis and loss of cilia induced by Mycoplasma hyopneumoniae in porcine tracheal organ cultures.

In vivo- and in vitro-grown Mycoplasma hyopneumoniae organisms were inoculated onto newborn piglet tracheal organ cultures to provide a model for interaction of this organism with ciliated respiratory epithelium. Ciliostasis and loss of cilia in tracheal rings were induced by M. hyopneumoniae grown in vivo and with low-passage cultures when grown in vitro. Levels of calmodulin or dehydrogenase enzymes in tracheal ring epithelium were not altered even though ciliostasis and loss of cilia induced by M. hyopneumoniae were extensive. The capacity for inducing epithelial damage diminished with in vitro passage of the organism. Attempts to induce higher-passage cultures to attach to cilia, cause ciliostasis, or cause ciliary damage by supplementation of mycoplasmal medium with porcine lung extract failed. Epithelial damage induced by M. hyopneumoniae in tracheal rings was averted by using porcine immune serum or by separating the organisms from ciliated epithelium with a 0.1-microns-pore-size membrane. Attachment, or at least close association, of M. hyopneumoniae to ciliated epithelium appeared to be necessary to induce ciliostasis and loss of cilia in this model.

[A cytogenetic study of the Chinese hamster lung cell line V-79 infected with Mycoplasma arginini and decontaminated using ciprofloxacin]. / Tsitogeneticheskoe issledovanie linii kletok legkogo kitaiskogo khomiachka V-79, infitsirovannoi mikoplazmoi Mycoplasma arginini i dekontaminirovannoi s pomoshch'iu tsiprofloksatsina.

Karyotypic variability has been studied in a line of the Chinese hamster cells artificially contaminated with Mycoplasma arginini. The contaminated cultures differed from mycoplasma-free cells in cell distribution for chromosome number. The frequency of cells with modal chromosome number 21 decreased, while that of cells with 20 chromosomes increased. Decontamination of cell culture with ciprofloxacin (10 mg/ml) and a subsequent cultivation of cell in the antibiotic-free medium did not restore the original cell distribution for chromosome number. In-53--131 days after infection, the increased frequency of chromosomal aberrations was registered. Eradication of Mycoplasma by ciprofloxacin and a long-term cultivation in antibiotic-free medium restored the frequency of chromosomal aberrations to the control level corresponding to mycoplasma-free cultures. In contaminated cultures no cyto- or genotoxic effect of ciprofloxacin was observed. These data, together with the previous ones, enable the authors to recommend ciprofloxacin to make cell cultures free from mycoplasmas with minimal risk to change the original properties of cell lines.

[Nonspecific immunotherapy using 2,4,6-trimethylhydroxypyrimidine (OPM-111) in Mycoplasma arthritidis infection in mice]. / Nespetsificheskaia immunoterapiia 2,4,6-trimetiloksipirimidinom (OPM-111) pri infektsii myshei Mycoplasma arthritidis.

Intraperitoneal infection of mice with M. anthritidis resulted in development of necrotic lesions in the omentum, mesentery and perirenal fatty tissue. The effect of 2,4,6-trimethyloxypyrimidine on such affections was studied. It was shown that the drug had immunomodulating properties and prevented development of lesions caused by Mycoplasma.

Diagnosis of genital Mycoplasma and Ureaplasma infections.

Genital Mycoplasma and Ureaplasma have been implicated in pelvic inflammatory disease, puerperal infections, septic abortions, low birth weight, nongonococcal urethritis and prostatitis as well as spontaneous abortion and infertility. An unequivocal diagnosis of infection with these organisms can be made only after properly obtained specimens have been evaluated with the use of selective cultures.

PIP: The fastidious growth requirements of mycoplasmas and ureaplasmas necessitated development of special growth media for them. The 1st mycoplasma was isolated from humans in 1937, and in 1954 a previously unknown mycoplasma was isolated from men with nonspecific urethritis. This organism, Ureaplasma urealyticum, is found most frequently in the genitourinary tract, followed by Mycoplasma hominus. M. fermentans and other mycoplasmas are isolated only rarely. Mycoplasmas and ureaplasmas have been implicated in pelvic inflammatory disease, puerperal infection, septic abortion, low birth weight, nongonococcal urethritis, and prostatisis, as well as spontaneous abortion and infertility, but there are no clinical symptoms pathognomonic of these infections. In spite of clinical suggestions of Mycoplasma or Ureaplasma infection, only a properly obtained specimen evaluatd with the use of selective cultures can lead to unequivocal diagnosis. The cultural characteristics and hence diagnostic procedures for Mycoplasma and Ureaplasma are quite different. Sterile calcium alginate swabs are used for obtaining urethral specimens, while sterile cotton swabs can be used for prostatic or vaginal secretions or semen. The swab should not touch antiseptic solutions, creams, or jellies, and the specimen must not dry out. Urine, if cultured, is best examined after centrifugattion at 600 g. Several different transport media are available. Optimally the specimen should be taken directly to the laboratory and subcultured on arrival. The metabolic activity of Mycoplasmas and Ureaplasmas is used in their detection. A phenol red indicator is added to the medium and the color change to or from yellow to pink indicates metabolic change. The growth medium is supplemented with glucose and phenol red for M. fermentans and arginine and phenol red for M. hominis. After color change is observed, the growth medium is subcultured on solid medium, which is obtained by adding .6-.8% Noble agar to the growth medium. Colonies develop best in an atmosphere of 95% N2 and 5% CO2 and reach approximately 200-300 mcm in diameter. They have a fried-egg appearance. Staining with Dienes stain, use of specific antisera, or incident light fluorescence microscopy are used for identification of the classic mycoplasmas. To isolate ureaplasmas, the specimen is transferred on arrival in the laboratory to urease color test broth U9C. During incubation the presence of Ureaplasma induces a rapid color change usually observable in 24-48 hours. A subculture should be done on fresh U9C broth media and on agar media once a color change is observed. Serologic tests for detection of antibodies to mycoplasmas and ureaplasmas are still in the developmental stage.