Abortion in cattle due to infection with Staphylococcus lugdunensis

Abstract

An aborted fetus of 7 months gestation, the associated placenta, and a single blood sample from the dam were submitted for diagnostic investigation to the diagnostic laboratory of the Lombardy and Emilia-Romagna Experimental Zooprophylactic Institute in Parma, Italy. The serum was negative for Neospora caninum, Coxiella burnetii, Chlamydophila abortus, Bovine herpesvirus 1 (BHV-1), Bovine viral diarrhea virus (BVDV), Brucella abortus, and Brucella melitensis. Fetal tissues and placental cotyledons were pooled and tested by polymerase chain reaction (PCR) for the presence of BHV-1, Bovine herpesvirus 4, BVDV, N. caninum, C. burnetii, Chlamydophila spp., Schmallemberg virus, and Leptospira interrogans. All PCR assays were negative. Bacteriological examinations performed on the fetal organs revealed a pure growth of Staphylococcus lugdunensis in all organs cultured. In human beings, S. lugdunensis is responsible for community-acquired and nosocomial infections, in both immunocompetent and immunocompromised patients. In veterinary medicine, the pathogenic potential of S. lugdunensis has not been fully investigated. The incidence of S. lugdunensis is regarded as being underreported because it could be easily misidentified as Staphylococcus aureus. The current report documents the ability of S. lugdunensis to cause abortion in cattle, indicating the need for accurate diagnostic procedures to identify this emerging and zoonotic pathogen whose incidence is likely underestimated in both human and veterinary medicine.

Abortion in cattle is defined as loss of the fetus occurring between 42 and 260 days of gestation. ⁷ The cause of this event is often difficult to identify, and the results of laboratory investigations reported in the literature suggest that only in approximately 35% of cases is it possible to ascertain the etiological agent. ⁴ Most bacteria causing bovine abortion are opportunistic pathogens. Trueperella pyogenes and Bacillus spp. followed by Escherichia coli, Histophilus somni, Pasteurella spp., Listeria monocytogenes, Staphylococcus spp., Streptococcus spp., and any other bacteria that can find their way into the bloodstream can act as opportunistic pathogens and cause sporadic abortion. Such opportunists are associated with abortions at any period of gestation, even though most are associated with late second to third trimester abortions. ¹⁵ In the current report, a case of abortion in cattle caused by Staphylococcus lugdunensis, an emerging pathogen in human beings ¹⁰ and companion animals,^22,24 is described.

Sporadic abortions were reported on a farm in the Province of Parma (northern Italy). The farm was a commercial dairy, consisting of approximately 500 lactating Holstein–Friesian cows. The abortion rate had been 3.8% over the preceding 12 months, and no laboratory diagnostics had been performed on those cases. In May 2013, a fetus of 7 months’ gestation, the associated placenta, and a blood sample of the cow were submitted for routine diagnostic investigation to the diagnostic laboratory of the Lombardy and Emilia-Romagna Experimental Zooprophylactic Institute in Parma, Italy.

The serum was negative for Neospora caninum, Coxiella burnetii, Chlamydophila abortus, Bovine herpesvirus 1 (BHV-1), Bovine viral diarrhea virus (BVDV), Brucella abortus, and Brucella melitensis. The veterinary practitioner who assisted the herd reported that no postabortion complications were observed in the dam.

The fetus presented minimal autolysis, moderate cutaneous edema, increased presence of fluids in the abdominal and thoracic cavities, moderately swollen spleen and liver, and complete atelectasis of the lungs. No gross lesions were apparent on the placenta.

Specimens of the brain, lung, heart, liver, spleen, kidney, abomasum content, and placental cotyledons were pooled and homogenized with sterile scissors for subsequent use in a number of polymerase chain reaction (PCR) assays. Pooling was a convenient compromise for fetal PCR diagnostics that broadened the spectrum of organs that could be tested while saving resources. Nucleic acids from the pool were extracted using commercial kits specific for DNA ^a and RNA ^b following manufacturers’ instructions. End-point PCRs were used to detect BHV-1, ⁸ Bovine herpesvirus 4, ⁸ BVDV, ¹⁸ N. caninum, ²⁰ C. burnetii, ² and Chlamydophila spp. ²³ A real-time PCR was used for Leptospira interrogans. ²⁷ Specimens of the brain, spleen, and thymus were also pool-tested for the presence of Schmallemberg virus by real-time PCR. ¹⁴ All PCR assays were negative.

Lung, liver, spleen, kidney, and the content of the abomasum were aseptically collected and cultured on bovine blood agar, ^c Gassner agar, ^d and Brucella agar ^e under aerobic and increased (10%) CO₂ incubation at 37°C. A pure culture appeared on the bovine blood agar ^c plates from all cultured organs and the abomasum content after 24 hr of incubation. After 48 hr, colonies were white, round, smooth, glistening, and 2–3 mm in diameter. A complete zone of hemolysis of approximately 1 mm was evident around the colonies. No growth was observed after 48 hr of incubation on Gassner agar ^d and after 10 days of incubation on Brucella agar. ^e Gram staining revealed the organism to consist of Gram-positive cocci. The bacterial isolate was catalase positive, oxidase negative, slide-coagulase positive, and tube-coagulase negative. A commercial identification kit ^f gave a profile of 6312150, which corresponded to S. lugdunensis with 97.3% identity. A second commercial identification system, ^g which includes 43 biochemical tests, was used as a confirmatory test. The second system gave a code of 010002002623231, corresponding to S. lugdunensis with 99% probability. No unexpected test results were obtained, and no supplementary tests were required for the definitive identification of the isolate.

Antimicrobial susceptibility was assessed by Kirby–Bauer disk diffusion according to the Clinical and Laboratory Standards Institute. ⁶ The strain was resistant to sulfadiazine, but sensitive to amoxicillin–clavulanic acid, ampicillin, ceftiofur, cloxacillin, colistin, enrofloxacin, florfenicol, oxacillin, penicillin, trimethoprim–sulfamethoxazole, tetracycline, thiamphenicol, tiamulin, and tylosin.

Staphylococcus lugdunensis is a coagulase-negative staphylococcus (CNS) that was first described in 1988. ¹¹ The organism is a commensal of the human skin more frequently found on the lower part of the body and the extremities, particularly in the moist areas such as the groin, perineum, and the large toenail. ³ Although CNS are generally considered nonpathogenic commensal in immunocompetent subjects, S. lugdunensis is an important exception being responsible for community-acquired and nosocomial infections, ¹⁰ in both immunocompetent^5,13,16,28 and immunocompromised patients.^13,21

In human beings, S. lugdunensis has been described as a possible cause of skin and soft tissue infection, prosthetic and naive valves endocarditis, bone and joint infections, and bacteremia. ¹⁰ The organism has rarely been isolated in peritonitis, and central nervous system, oral, ocular, and urinary tract infections. ¹⁰ Staphylococcus lugdunensis has also been associated with endometritis, ¹ pyomyoma, ²⁶ and, in 2013, its detection in amniotic fluids collected during cesarean sections was reported, although no postoperative infection or neonatal complications were observed. ¹⁹

In veterinary medicine, the pathogenic potential of S. lugdunensis has not been fully investigated. A retrospective case control study of S. lugdunensis infections in small companion animals suggested that S. lugdunensis is more often isolated from deep tissues and wounds than from superficial infections, therefore, it should be considered a potential pathogen. ²⁴ Staphylococcus lugdunensis was also isolated from a blood sample of a dog with endocarditis, but a role in the pathogenesis of the disease remains uncertain as the dog was also positive for Bartonella spp. by PCR, and possible contamination of the blood sample by S. lugdunensis through the skin could not be ruled out. ²² Staphylococcus lugdunensis was also shown to cause subcutaneous abscesses, peritonitis, and osteomyelitis in mice^9,17,25 and rabbits ¹² used in experimental studies.

The incidence of S. lugdunensis is presumed to be underreported in both human and veterinary medicine as it can be easily misidentified as Staphylococcus aureus, which shares the same colony morphology, hemolytic activity, and the ability to agglutinate latex particles coated with fibrinogen. ²⁹ The current report documents the ability of S. lugdunensis to cause abortion in cattle, underlining the need for accurate diagnostic procedures to correctly identify this emerging and zoonotic pathogen whose incidence is likely underestimated.