Identification of a novel Hemoplasma species from pigs in Zhejiang province, China.

Hemoplasmas belong to Mycoplasmataceae (Mollicutes: Mycoplasmatales) and are able to infect a broad range of mammalian species. We investigated prevalence of hemotropic mycoplasma species in pig farms in the region of Zhejiang by a PCR scheme using universal primers targeting 16S rRNA and RNase P RNA gene (rnpB). Representative positive samples from different farms were selected for sequencing of 16S rRNA and the 219bp rnpB gene fragments for phylogenetic analysis. Sequencing analysis of PCR products from first samples identified a novel hemoplasma species present in several pig farms in the region with highest nucleotide identity of 92% to Candidatus Mycoplasma turicensis. A duplex PCR assay was then designed for differential detection of the novel hemoplasma from Mycoplasma parvum/M. suis in field samples. Of 324 blood samples from clinically healthy pigs, 26.5% was positive for this novel hemoplasma species and 50% positive for M. suis/M. parvum, indicating that the novel hemotropic mycoplasma species were of considerably high prevalence in Zhejiang province, China.

Development and clinical application of an InvaderPlus® assay for the detection of genital mycoplasmas.

We developed a PCR-based assay involving Invader® technology for detection of the genital mycoplasmas of Mycoplasma genitalium, Mycoplasma hominis, Ureaplasma urealyticum, and Ureaplasma parvum. We compared its performance with that of a PCR-microtiter plate hybridization assay, which we developed previously, in detecting genital mycoplasmas in first-voided urine (FVU) specimens from men with non-gonococcal urethritis. The tests targeting each of the genital mycoplasmas were specific for the respective species and could detect as few as 10 copies of the plasmids containing the target genes of each of the genital mycoplasmas per reaction. The assay using the InvaderPlus® method (InvaderPlus® assay) showed very similar performance to that of the PCR-microtiter plate hybridization assay for detecting the genital mycoplasmas in the FVU specimens. In addition, the PCR and endonuclease reaction in the InvaderPlus® assay were carried out simultaneously in one procedure, thus simplifying the assay, leading to time- and labor-savings and a decrease in the risk of specimen contamination. The InvaderPlus® assay could be useful in diagnosing genitourinary tract infections caused by the genital mycoplasmas.

Treatment of genital mycoplasma in colonized pregnant women in late pregnancy is associated with a lower rate of premature labour and neonatal complications.

Mycoplasma hominis and Ureaplasma spp. may colonize the human genital tract and have been associated with adverse pregnancy outcomes such as preterm labour and preterm premature rupture of membranes. However, as these bacteria can reside in the normal vaginal flora, there are controversies regarding their true role during pregnancy and so the need to treat these organisms. We therefore conducted a retrospective analysis to evaluate the treatment of genital mycoplasma in 5377 pregnant patients showing symptoms of potential obstetric complications at 25-37 weeks of gestation. Women presenting with symptoms were routinely screened by culture for the presence of these bacteria and treated with clindamycin when positive. Compared with uninfected untreated patients, women treated for genital mycoplasma demonstrated lower rates of premature labour. Indeed preterm birth rates were, respectively, 40.9% and 37.7% in women colonized with Ureaplasma spp. and M. hominis, compared with 44.1% in uncolonized women (Ureaplasma spp., p 0.024; M. hominis, p 0.001). Moreover, a reduction of neonatal complications rates was observed, with 10.9% of newborns developing respiratory diseases in case of Ureaplasma spp. colonization and 5.9% in the presence of M. hominis, compared with 12.8% in the absence of those bacteria (Ureaplasma spp., p 0.050; M. hominis, p <0.001). Microbiological screening of Ureaplasma spp. and/or M. hominis and pre-emptive antibiotic therapy of symptomatic pregnant women in late pregnancy might represent a beneficial strategy to reduce premature labour and neonatal complications.

Antimicrobial susceptibility patterns of Ureaplasma species and Mycoplasma hominis in pregnant women.

BACKGROUND: Genital mycoplasmas colonise up to 80% of sexually mature women and may invade the amniotic cavity during pregnancy and cause complications. Tetracyclines and fluoroquinolones are contraindicated in pregnancy and erythromycin is often used to treat patients. However, increasing resistance to common antimicrobial agents is widely reported. The purpose of this study was to investigate antimicrobial susceptibility patterns of genital mycoplasmas in pregnant women. METHODS: Self-collected vaginal swabs were obtained from 96 pregnant women attending an antenatal clinic in Gauteng, South Africa. Specimens were screened with the Mycofast Revolution assay for the presence of Ureaplasma species and Mycoplasma hominis. The antimicrobial susceptibility to levofloxacin, moxifloxacin, erythromycin, clindamycin and tetracycline were determined at various breakpoints. A multiplex polymerase chain reaction assay was used to speciate Ureaplasma positive specimens as either U. parvum or U. urealyticum. RESULTS: Seventy-six percent (73/96) of specimens contained Ureaplasma spp., while 39.7% (29/73) of Ureaplasma positive specimens were also positive for M. hominis. Susceptibilities of Ureaplasma spp. to levofloxacin and moxifloxacin were 59% (26/44) and 98% (43/44) respectively. Mixed isolates (Ureaplasma species and M. hominis) were highly resistant to erythromycin and tetracycline (both 97% resistance). Resistance of Ureaplasma spp. to erythromycin was 80% (35/44) and tetracycline resistance was detected in 73% (32/44) of Ureaplasma spp. Speciation indicated that U. parvum was the predominant Ureaplasma spp. conferring antimicrobial resistance. CONCLUSIONS: Treatment options for genital mycoplasma infections are becoming limited. More elaborative studies are needed to elucidate the diverse antimicrobial susceptibility patterns found in this study when compared to similar studies. To prevent complications in pregnant women, the foetus and the neonate, routine screening for the presence of genital mycoplasmas is recommended. In addition, it is recommended that antimicrobial susceptibility patterns are determined.

The prevalence of six species of Mycoplasmataceae in an HIV/AIDS population in Jiangsu Province, China.

This study employed culture and polymerase chain reaction (PCR) to examine the prevalence of Ureaplasma urealyticum, Mycoplasma hominis, Mycoplasma genitalium, Mycoplasma fermentans, Mycoplasma penetrans and Mycoplasma pirum in 210 HIV/AIDS patients, 455 sexually transmitted infection (STI) clinic attendees and 245 healthy volunteers from first-void urine specimens for men and endocervical swabs for women. U. urealyticum and M. hominis were detected in 107 (51.0%) and 69 (32.9%) patients in the HIV/AIDS group. At least one of the other four organisms was detected in 34 (16.2%) HIV/AIDS patients, 29 (6.4%) STI clinic attendees and six (2.5%) healthy volunteers. This study showed that U. urealyticum, M. hominis and M. fermentans were significantly more prevalent in HIV/AIDS patients, as were other mycoplasmas. Our results suggest a possible role for co-infection.

Evaluation of Seeplex® STD6 ACE Detection kit for the diagnosis of six bacterial sexually transmitted infections.

Traditionally, the diagnosis of bacterial sexually transmitted infection (STI) has been dependent on the isolation of the causative pathogens by culturing endocervical or urethral swab specimens on selective media. While such procedures typically provide excellent diagnostic accuracy, they are often time-consuming and expensive. A multiplex polymerase chain reaction (PCR) assay, based on a semi-automated detection system, was evaluated for the detection of six STI causative organisms. The Seeplex(®) STD6 ACE (auto-capillary electrophoresis) Detection assay employed six pairs of dual priming oligonucleotide (DPO™) primers specifically targeted to unique genes of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium, Ureaplasma urealyticum, Mycoplasma hominis, and Trichomonas vaginalis. A total of 739 specimens (304 cervical swabs and 435 urine samples) collected for 4 months were tested, and results were compared to those obtained with a combined monoplex PCR. The concordance between the multiplex PCR and monoplex PCR assay was 100% for both sensitivity and specificity. We also tested for the presence of two pathogenic bacteria (C. trachomatis and N. gonorrhoeae) and compared the results obtained with the multiplex PCR and BD ProbeTec duplex strand displacement amplification (SDA). The results of the multiplex PCR and duplex SDA were 99.7% concordant for C. trachomatis and 100% concordant for N. gonorrhoeae. The multiplex PCR assay using the Seeplex(®) STD6 ACE Detection kit proved to be a novel cost-effective and fast diagnostic tool with high sensitivity and specificity for the simultaneous detection of six STI pathogens.

Comparison of multiplex PCR assay with culture for detection of genital mycoplasmas.

Ureaplasma, spp. Mycoplasma genitalium, and Mycoplasma hominis are associated with infection of the genitourinary tract, reproductive failure, and neonatal morbidity and mortality. We have developed a multiplex PCR for the detection of these Mycoplasmatales in a single amplification reaction. The analytical sensitivities of this assay were 10.8, 10.8, and 8.8 CFU for each organism, respectively. This multiplex PCR was compared to culture on 26 cervical swabs, 2 vaginal swabs, 4 female urine specimens, 49 semen samples, 2 male urine specimens, and 1 nonspecified sample. A total of 21 specimens were culture positive (25%); 17 of these were PCR positive. An additional 11 specimens were PCR positive but culture negative. Of the 21 culture-positive specimens, 17 (81%) grew Ureaplasma spp. and 4 (19%) grew Mycoplasma spp. Of the 28 PCR-positive specimens, Ureaplasma spp. was detected in 23 (82%), M. hominis was detected in 3 (11%), and both were detected in 2 (7%). In a confirmatory analysis, all samples were tested by amplification of a second target of the ureaplasma genome. True-positive cases were defined as a positive result by culture or by both amplification assays. The multiplex PCR detected organisms in 26 of the 30 true-positive specimens, as well as in 2 other specimens. Based on a 36% prevalence of infection, the sensitivity, specificity, and positive and negative predictive values of multiplex PCR analyses were 87, 96, 94, and 93%, respectively. Multiplex PCR offers a rapid, sensitive, and easy method to detect genital mycoplasmas.

Mycoplasmal antibodies as determined with an enzyme-linked immunosorbent assay, in tubal factor infertility.

A total of 81 infertile women, who had been referred for diagnostic loparoscopy, were tested for the presence of antibodies to Mycoplasma hominis and T-mycoplasma. Out of 81, 30 had tubal adhesions and 51 had unilateral/bilateral tubal blockage. Antibodies to M. hominis were found in 21/30 (70%) and 14/51 (27.45%) women, antibodies to T-mycoplasma in 12/20 (40% and 39/51 (76.47%) women with tubal disorder. In a control group of 40 pregnant women, antibodies to the same two organisms occurred in 10% and 32.5%. Antibodies to M. hominis and T-mycoplasma were significantly (P < 0.001) more common in women with tubal disorder. Our results confirm the important role of M. hominis and T-mycoplasma in the aetiology of tubal infertility.

Spiroplasmas: infectious agents of plants, arthropods and vertebrates.

The spiroplasmas are mollicutes characterized by motility and helical morphology. They were discovered through studies on corn stunt and citrus stubborn diseases. The stubborn agent was the first mollicute of plant origin to be obtained in culture and the first cultured mollicute to possess a helical morphology. The citrus pathogen has been known as Spiroplasma citri since 1973. The corn stunt agent was cultured in 1975 and fully characterized as Spiroplasma kunkelii by 1986. The third and only other phytopathogenic spiroplasma is Spiroplasma phoeniceum, cultured from naturally infected periwinkle plants in Syria and described in 1986. These three spiroplasmas are restricted to the phloem sievetubes of the infected plants and are transmitted from plant by various phloem feeding leafhopper vectors in which the spiroplasmas multiply. Following the pioneering work on S. citri and S. kunkelii, close to fifty other spiroplasma species or proposed species have been discovered. All spiroplasmas have been isolated from insects, ticks and plants. Insects are particularly rich sources of spiroplasmas. Some insect-derived spiroplasmas are entomopathogens. S. melliferum and S. apis are honey bee pathogens. They cross the insect-gut barrier and reach the hemolymph, where they multiply abundantly and kill the bee. Spiroplasma floricola is the agent of lethargy disease of Melolontha melolontha (cockchafer). Spiroplasma poulsonii infects the neotropical species of Drosophila, is transmitted transovarially and kills the male progeny of an infected female fly, hence the name sex ratio spiroplasma. Some insect-derived spiroplasmas are also found on plant (flower) surfaces. For instance, S. apis was cultured from the surfaces of flowers growing in the vicinity of affected beehives. This suggests that the plant surface spiroplasmas are deposited on these surfaces by contaminated insects. Many insect spiroplasmas are not pathogenic, are often restricted to the gut and may be regarded as mutualists or incidental commensals. Of the three known tick spiroplasmas, only Spiroplasma mirum obtained from rabbit ticks is pathogenic to the vertebrate animal (chick embryo, new-born rodents, adult rabbit), but only upon experimental inoculation of the spiroplasma. Strain SMCA induces high incidence of cataracts in new born rodents. With strain GT-48 no cataracts are observed, but fatal encephalitis occurs. Spiral membranous inclusions resembling spiroplasmas have been seen in brain biopsies taken from patients with Creutzfeldt-Jakob disease. However, failure to detect spiroplasmas by serology and culture points to the absence of spiroplasmal involvement in spongiform encephalopathies. Transposon Tn 4001 mutagenesis has been applied for the first time to Spiroplasma citri, and pathogenicity can now be studied at the genetic level. One Tn 4001 mutant does not multiply in the leafhoppers and is, therefore, not transmitted to the plant. Another mutant multiplies well in the plant and is transmitted to the plant, where it reaches high titers, but without inducing symptoms in the plant. In this non-phytopathogenic mutant, Tn 4001 is inserted in the spiroplasmal fructose operon, and the mutant is unable to use fructose. Finally, to study involvement of spiroplasmal motility in pathogenicity, a non-motile mutant has been obtained. Motility was restored by complementation with the wild type genes. This is the first time that successful complementation has been reported, not only in the spiroplasmas but in the mollicutes in general. Undoubtedly, studies on pathogenicity have entered a new era.

[The effect of Mycoplasma contamination and decontamination with ciprofloxacin on the karyotypic structure of the Chinese hamster V-79 lung cell line]. / Vliianie mikoplazmennoi kontaminatsii i dekontaminatsii s pomoshch'iu tsiprofloksatsina na kariotipicheskuiu strukturu kletochnoi linii legkogo kitaiskogo khomiachka V-79.

Karyotypic variability was investigated for the Chinese hamster lung cell line V-79, infected (contaminated) with a mycoplasma Acholeplasma laidlawii A. The mycoplasmal contamination did not affect cell distribution for the chromosome number. However, 30-70 days following cell culture contamination the increase in chromosomal aberrations was observed in the contaminated cell line primarily at the expense of chromosomal breaks. The following cyprofloxacin treatment of the culture resulted in mycoplasmal elimination and decrease in the frequency of chromosomal aberrations up to the control level. The analysis of G-banded chromosomes showed no significant differences in karyotypes of originally non-infected cells and cells after decontamination. The karyotypic variability, induced by mycoplasmal infection in V-79 cell line, differed from that in the muntjac skin fibroblast cell line, the latter being described elsewhere. A predominant type of chromosomal variability in V-79 cell line are chromosomal breaks, whereas in the muntjac fibroblast cell line dicentrics (telomeric fusions) were primarily observed, possible explanations of these differences being discussed. This may be presumably due to differences in karyotypic structure of these two cell lines, and to their different adaptation to culture conditions.

The nasal mycoplasmal flora of healthy calves and cows.

The nasal mycoplasmal flora of 270 healthy cows from 27 herds in the Netherlands and 35 healthy calves from 7 of these herds was examined. Various methods for isolating mycoplasmas were compared. The prevalence of the various species was as follows: Ureaplasma diversum in 3 (9%) calves; Mycoplasma dispar in 14 (40%) calves; M. bovis in 1 (3%) calf; M. bovirhinis in 23 (66%) calves and 16 (6%) cows; M. bovoculi in 8 (23%) calves and 53 (20%) cows; M. canis in 1 (3%) calf; M. equirhinis in 2 (1%) cows; M. conjunctivae in 2 (1%) cows; Acholeplasma laidlawii in 1 (3%) calf and 3 (1%) cows; and A. axanthum in 7 (3%) cows. The noses of healthy calves were less frequently colonized by the pathogenic species U. diversum and contained fewer U. diversum and M. dispar organisms than the noses of pneumonic calves. We concluded that the mycoplasmal flora of calves and healthy cows was quite different and also that cows play only a minor role in the epidemiology of pathogenic mycoplasma species of calves in the Netherlands.

Clinical and biological characteristics of Ureaplasma urealyticum induced polyarthritis in a patient with common variable hypogammaglobulinaemia.

Persistent infectious polyarthritis caused by Ureaplasma urealyticum in a patient with common variable hypogammaglobulinaemia is described. The patient developed a symmetrical, destructive polyarthritis and tenosynovitis associated with a markedly depressed synovial fluid glucose concentration and characteristic soft tissue abscesses. The ureaplasma organism developed resistance to multiple antibiotics and persisted for five years. The organism was identified repeatedly in many joints by culture, confirmed by DNA hybridisation, and mycoplasma-like structures were shown in synovial tissues by electron microscopy.

[Ureaplasma urealyticum--a new problem pathogen in neonatology]. / Ureaplasma urealyticum--ein neuer Problemkeim in der Neonatologie.

Some previous studies showed that Ureaplasma urealyticum is the most common germ that appears in the birthway of pregnant women and which is also frequently found in skin swabs and secretions of newborn and premature babies. The colonization of pregnant women by Ureaplasma urealyticum makes a premature birth more likely. Another factor of risk for a premature infant is a premature rupture of membranes for more than 24 hours which also makes an infection possible. There exists an association between pulmonary infection by Ureaplasma urealyticum and the development of a bronchopulmonary dysplasia especially for premature babies. According to our observations acute exacerbations of severe pneumonia can appear even after month. An attempt of therapy of pulmonary infection should be undertaken with erythromycin, if sensitive serotypes are present. In the case of erythromycin resistance chloramphenicol can be used but only under frequent controls of blood levels. We were able to observe rapid improvements with this effective therapy.

Survival of feline mycoplasmas in urine.

The effects of length of incubation and urine osmolality on the survival of feline mycoplasmas and ureaplasmas and representative gram-positive and gram-negative bacteria in synthetic urine which approximated the osmolality of normal cat urine were investigated. Both Escherichia coli and Staphylococcus aureus withstood the effects of increasing osmotic pressure. In the most concentrated urine, significant decreases (P less than 0.001) in CFU were observed for E. coli at exposure times of 30 min and longer. S. aureus was not affected by longer exposure or increased osmotic strength. Both Mycoplasma felis and Mycoplasma gateae were affected adversely by longer exposure times and high osmotic strength (P less than 0.001). A Ureaplasma sp. was not adversely affected except at very high (greater than or equal to 2,980 mosM) osmotic strengths or after prolonged incubation (120 min) at relatively high (1,976 mosM) osmotic strengths (P less than 0.001). The failure of both M. felis and M. gateae to survive under osmotic conditions present in normal feline urine suggests that it is unlikely that these mycoplasmas are involved in urinary disorders in cats.

[Infections by Mycoplasma hominis and Ureaplasma urealytycum in vaginal discharge. Incidence and treatment]. / Infections à Mycoplasmes hominis et Ureaplasma urealytycum dans les écoulements vaginaux. Fréquence et traitements.

This report summarizes the studies carried out on 100 patients who were examined for vaginal discharge. The following observations were made: Mycoplasma hominis in 13.59% of the cases, Ureaplasma urealytycum in 8.73% of the cases, that is, the presence of at least one of these microorganisms in 22.32 of the cases. All these infections were treated by tetracyclines administered in appropriate doses.

[Detection of Mycoplasma infection in the postpartum period]. / Vyiavlenie mikoplazmennoi infektsii u zhenshchin v poslerodovom periode.

The comparative results of microbiological and serological testing of parturients with uneventful and complicated course of the postpartum period were presented. The data justified the etiological role of M. hominis and U. urealyticum in the development of postpartum complications.