Mycoplasma miroungirhinis sp. nov. and Mycoplasma miroungigenitalium sp. nov., isolated from northern elephant seals (Mirounga angustirostris), Mycoplasma phocoenae sp. nov., isolated from harbour porpoise (Phocoena phocoena), and Mycoplasma phocoeninasale sp. nov., isolated from harbour porpoise and California sea lions (Zalophus californianus).

Seven novel independent strains of Mycoplasma species were isolated from northern elephant seals (ES2806-NAST, ES2806-GENT, ES3157-GEN-MYC and ES3225-GEN-MYC), a harbour porpoise (C264-GENT and C264-NAST), and a California sea lion (CSL7498). These strains were phenotypically and genetically characterized and compared to the known Mycoplasma species. Four strains (C264-GENT, C264-NAST, CSL7498 and ES2806-NAST) hydrolysed arginine but not urea and did not produce acid from carbohydrates. Strains ES2806-GENT, ES3157-GEN-MYC and ES3225-GEN-MYC did not produced acid from carbohydrates and did not hydrolyse arginine or urea; hence, it is assumed that organic acids are used as the energy source for them. All were isolated and propagated in ambient air supplemented with 5±1 % CO2 at +35-37 °C using either SP4 or PPLO medium. Colonies on solid medium showed a typical fried-egg appearance and transmission electron microscopy revealed a typical mycoplasma cellular morphology. The complete genomes were sequenced for all type strains. Average nucleotide and amino acid identity analyses showed that these novel strains were distant from the phylogenetically closely related Mycoplasma species. Based on these data, we propose four novel species of the genus Mycoplasma, for which the name Mycoplasma miroungirhinis sp. nov. is proposed with the type strain ES2806-NAST (=NCTC 14430T=DSM 110945T), Mycoplasma miroungigenitalium sp. nov. is proposed with the type strain ES2806-GENT (=NCTC 14429T=DSM 110944T) and representative strains ES3157-GEN-MYC and ES3225-GEN-MYC, Mycoplasma phocoenae sp. nov. is proposed with the type strain C264-GENT (=NCTC 14344T=DSM 110687T) and Mycoplasma phocoeninasale sp. nov. is proposed with the type strain C264-NAST (=NCTC 14343T=DSM 110688T) and representative strain CSL7498. The genome G+C contents are 24.06, 30.09, 28.49 and 29.05% and the complete genome sizes are 779 550, 815 486, 693 115, and 776 009 bp for strains ES2806-NAST, ES2806-GENT, C264-GENT and C264-NAST, respectively.

STREPTOCOCCUS PHOCAE IN MARINE MAMMALS OF NORTHEASTERN PACIFIC AND ARCTIC CANADA: A RETROSPECTIVE ANALYSIS OF 85 POSTMORTEM INVESTIGATIONS.

: Streptococcus phocae is a pathogen of marine mammals, although its pathogenicity remains poorly understood. Recovery of this bacterium from asymptomatic carriers suggests that it is an opportunistic pathogen. We investigated the role of S. phocae in naturally occurring disease and its significance as a pathogen based on postmortem investigations. Between 2007 and 2012, 1,696 whole carcasses, tissue samples, or both were submitted from the northeastern Pacific and Arctic Canada for diagnostic testing. Streptococcus phocae was cultured from phocids ( n=66), otariids ( n=12), harbor porpoises ( Phocoena phocoena; n=5), and sea otters ( Enhydra lutris; n=2). Pathologic manifestations of S. phocae-associated disease included localized, as well as systemic, inflammatory lesions with common findings of suppurative bronchopneumonia ( n=17) and bacteremia ( n=27). Lung lesions were frequently culture-positive for S. phocae, suggesting commensal colonization of the oropharynx with subsequent opportunistic infection of the respiratory tract during tissue injury, coinfection, immunosuppression, or other debilitating conditions. The presence of a positive spleen culture, and interpretations at necropsy and histopathology, were used to determine the presence of S. phocae bacteremia. Less frequent lesions that were culture positive for S. phocae included abscesses ( n=9), meningitis ( n=7), and cellulitis ( n=1). The majority of cases with S. phocae lesions featured pre-existing conditions that presumably contributed to some degree of debilitation or immunosuppression, including emaciation ( n=29), liver mercury accumulation ( n=29), trauma ( n=22), severe pulmonary or cardiovascular nematodiasis ( n=9), concurrent bacterial or viral infections ( n=8), or sarcocystosis ( n=6). These findings suggest that S. phocae could be characterized as an opportunistic pathogen, associated with debilitating conditions in stranded and rehabilitating marine mammals. Wildlife investigators can use these results to draw more definitive conclusions regarding positive S. phocae cultures during postmortem studies in marine mammals.

Population genetic structure of 4,12:a:- Salmonella enterica strains from harbor porpoises.

According to pulsed-field gel electrophoresis (PFGE) typing, 4,12:a:- Salmonella enterica isolates from harbor porpoises are highly diverse. However, porpoise isolates belong to only two multilocus sequence types within the eBurst group 18 (eBG18) genetic cluster, which also includes S. enterica serovars Bispebjerg and Abortusequi. Isolates of other, serologically similar serovars belong to unrelated eBGs. These assignments to eBGs were supported by eBG-specific sequences of the flagellar gene fliC.

Human exposures to marine Brucella isolated from a harbor porpoise - Maine, 2012.

On February 10, 2012, the Maine Center for Disease Control and Prevention (Maine CDC) was notified of a positive Brucella culture from a harbor porpoise (Phocoena phocoena) found on the coast of southern Maine. Maine CDC, in consultation with CDC, initiated an investigation of potential occupational exposures of staff members at university A and at diagnostic laboratories known to have handled samples from the porpoise. This report describes the results of that investigation. In humans, brucellosis can cause fever, sweats, headaches, back pains, physical weakness, and sometimes severe infections of the brain, bone, heart, liver, or spleen. Because staff members at university A did not use respiratory protection while handling the porpoise or its specimens, the four exposed staff members were advised to begin immediately a 3-week regimen of rifampicin and doxycycline for antimicrobial prophylaxis, conduct daily fever checks, be monitored for symptoms of acute febrile illness weekly, and have their serum tested for Brucella antibodies immediately and at regular intervals for 24 weeks after the last known exposure. As of June 26, none of the four persons had seroconverted or become ill. The potential for human infection and illness as well as the intensity, duration, and expense of the follow-up recommended for Brucella exposure highlights the need for facilities to develop standard protocols for preventing exposures during the handling of marine mammals, particularly during aerosol-generating procedures.

Brucella ceti infection in harbor porpoise (Phocoena phocoena).

We describe Brucella sp. infection and associated lesions in a harbor porpoise (Phocoena phocoena) found on the coast of Belgium. The infection was diagnosed by immunohistochemistry, transmission electron microscopy, and bacteriology, and the organism was identified as B. ceti. The infection's location in the porpoise raises questions of abortion and zoonotic risks.

Prevalence of a host-adapted group B Salmonella enterica in harbour porpoises (Phocoena phocoena) from the south-west coast of England.

A monophasic group B Salmonella enterica 4,12:a:- was first isolated in harbour porpoises (Phocoena phocoena) in Scotland in 1991. This paper reports the isolation of the same group B S enterica from harbour porpoise carcases found stranded along the Cornwall and Devon coastlines. Between 1991 and 2002, 80 harbour porpoises were submitted for postmortem examination and subjected to bacteriological examination under the UK Cetacean Strandings Investigation Programme. A total of 28 Salmonella isolates were recovered and subjected to several tests, including biochemical, molecular and serological analysis.

Mycotic otitis media in a harbour porpoise (Phocoena phocoena).

A 9-year-old by-caught harbour porpoise (Phocoena phocoena) was subject to routine post-mortem examination. Major findings included parasitic infection (Stenurus minor) of the left middle ear and severe mycotic infection of the right middle ear. The morphological appearance of the causative organism was consistent with Aspergillus fumigatus. These pathological findings were likely to have impaired echolocation in this animal and this may have contributed to the by-catch. The ears of marine mammals should be examined routinely during post-mortem investigations.

Regional differences in bacterial flora in harbour porpoises from the North Atlantic: environmental effects?

AIMS: Microbiological findings in harbour porpoises from different regions of the North Atlantic were compared. Results in animals from the North and Baltic Seas were evaluated over a period of 18 years for changes in the microbiological flora. METHODS AND RESULTS: Microbiological investigations were performed on 1429 organ samples from the lung, liver, kidney, spleen, intestine, and mesenteric lymph nodes from harbour porpoises of the German North and Baltic Seas, Greenlandic, Icelandic and Norwegian waters. A large variety of bacteria, including potentially pathogenic bacteria like Brucella sp., Clostridium perfringens, Escherichia coli, Erysipelothrix rhusiopathiae, beta-haemolytic streptococci and Staphylococcus aureus were isolated. Those bacteria were associated with bronchopneumonia, gastroenteritis, hepatitis, pyelonephritis, myocarditis and septicemia. CONCLUSIONS: Organs from animals originating from Greenlandic and Icelandic waters showed clearly less bacterial growth and fewer associated pathological lesions compared to animals from the German North and Baltic Seas and Norwegian waters. SIGNIFICANCE AND IMPACT OF THE STUDY: Differences in bacterial findings and associated lesions between harbour porpoises from the German North and Baltic Seas and animals from Greenlandic, Norwegian and Icelandic waters may result from higher stress due to anthropogenic activities such as chemical pollutants in the North and Baltic Seas.

Isolation of Mycobacterium mageritense from the lung of a harbor porpoise (Phocoena phocoena) with severe granulomatous lesions.

Post-mortem investigation of a harbor porpoise (Phocoena phocoena) found dead on the beach of the island of Vlieland, The Netherlands, revealed severe granulomatous changes in the right lung lobe. Ziehl Neelsen staining demonstrated relatively large acid-fast rods. Mycobacterial culture yielded a fast-growing mycobacterium, which was identified by molecular biological methods as Mycobacterium mageritense. Autolysis prevented histopathology. It was tentatively concluded that the granulomatous changes were the cause of porpoise's death and that M. mageritense was the causative agent. This is the first report of the isolation and molecular identification of this mycobacterium in a nonhuman animal species and the first association with the marine environment.

Occurrence of a monophasic strain of Salmonella group B isolated from cetaceans in England and Wales between 1990 and 2002.

Between September 1990 and December 2002, 511 cetacean carcasses stranded or caught in commercial fisheries in England and Wales were examined post mortem. Salmonella group B was isolated from 60 of 279 (21.51%) harbour porpoises (Phocoena phocoena), predominantly from lung tissue. Forty-three of the Salmonella group B isolates were subsequently serotyped and all found to have the antigenic structure O4,12:a:-. The annual proportion of harbour porpoises testing positive for Salmonella O4,12:a:- increased significantly from 6% in the early 1990s to 27% after 1999. The cause(s) of the increasing prevalence of Salmonella O4,12:a:- are not known, but may reflect natural variation in the epidemiological cycle of Salmonella O4,12:a:- in harbour porpoises. The probability of isolating Salmonella O4,12:a:- from harbour porpoises increased with age, suggesting that the mode of transmission is principally horizontal. There appeared to be a weak degree of seasonality in the probability of isolating Salmonella O4,12:a:- with a low proportion of cases in the months of April and May. Based on pathological findings from infected carcasses, Salmonella O4,12:a:- may be part of the normal commensal flora of the lungs of cetaceans with the potential to act as an opportunistic pathogen.

Increased blood interleukin-10 mRNA levels in diseased free-ranging harbor porpoises (Phocoena phocoena).

Harbor porpoises from the North and Baltic Seas exhibit a higher incidence of bacterial infections compared to whales from less polluted arctic waters. Toxicological analysis revealed an association between elevated body burdens of environmental contaminants, such as polychlorinated biphenyls (PCB) and polybrominated diphenyl ether (PBDE) and lymphoid depletion in thymus and spleen of these whales. However, it remains undetermined if changes in the immune system are primarily contaminant-induced or a sequel of infectious diseases and emaciation. The aim of the present study was to investigate changes of blood cytokine mRNA levels in healthy and diseased harbor porpoises. Therefore, 29 by-caught and stranded whales were necropsied and the health status was evaluated based upon main pathological findings. Furthermore, the degree of thymic atrophy and splenic depletion was histologically graded using a semi-quantitative scoring system. Gene expression of interleukin (IL)-2, IL-4, IL-6, IL-10, transforming growth factor-beta and tumor necrosis factor-alpha in the blood was measured by real time reverse transcription-polymerase chain reaction. Thymic atrophy and splenic depletion were correlated with an impairment of the animals' health status. Additionally, a marked up-regulation of IL-10 was predominately found in severely diseased whales with evidence of chronic bacterial infections. Furthermore, increased IL-10 levels were associated with splenic depletion. Other investigated cytokines were not significantly associated with the health status or lymphoid depletion, respectively. The present study indicated that lymphoid depletion represents a sequel of chronic infectious diseases in a portion of investigated harbor porpoises. Regarding this, expression of anti-inflammatory cytokines, such as IL-10 might represent a consequence of continuous stimulation of the immune system and induction of immunomodulatory mechanisms in this cetacean species.

Pathological findings in harbour porpoises (Phocoena phocoena) from Norwegian and Icelandic waters.

A study of 37 by-caught harbour porpoises from Icelandic and Norwegian waters showed that most were in good or moderate nutritional condition and none was severely emaciated. Mild infection with lungworms (Halocercus invaginatus, Pseudalius inflexus, Torynurus convolutus) was found in 84% of the Icelandic and 91% of the Norwegian animals, usually associated with bronchopneumonia which was rarely severe. Most (91%) of the animals had parasites in the stomach and intestine (Anisakis simplex, Contracaecum osculatum, Pholeter gastrophilus), and Campula oblonga was present in the liver and pancreas of 88 and 21%, respectively. Oesophagitis, gastritis, cholangitis, pericholangitis, pancreatitis and lymphadenitis were almost exclusively associated with parasitic infection and usually mild. Bacterial isolates were obtained from 50 to 55% of the animals but were not considered to be clinically significant. There was no indication of morbillivirus infection. Icelandic and Norwegian animals showed a thicker blubber layer and a lower incidence of severe lesions, especially in the respiratory tract, as compared with reports of by-caught animals from the Baltic Sea.

Bartonella henselae in porpoise blood.

We report detection of Bartonella henselae DNA in blood samples from 2 harbor porpoises (Phocoena phocoena). By using real-time polymerase chain reaction, we directly amplified Bartonella species DNA from blood of a harbor porpoise stranded along the northern North Carolina coast and from a pre-enrichment blood culture from a second harbor porpoise. The second porpoise was captured out of habitat (in a low-salinity canal along the northern North Carolina coast) and relocated back into the ocean. Subsequently, DNA was amplified by conventional polymerase chain reaction for DNA sequencing. The 16S-23S intergenic transcribed spacer region obtained from each porpoise was 99.8% similar to that of B. henselae strain San Antonio 2 (SA2), whereas both heme-binding phage-associated pap31 gene sequences were 100% homologous to that of B. henselae SA2. Currently, the geographic distribution, mode of transmission, reservoir potential, and pathogenicity of bloodborne Bartonella species in porpoises have not been determined.