Brucellosis is a zoonotic disease of worldwide distribution. It negatively impacts human health, and animal production and economy by significant loss . Brucellosis may become chronic causing severe osteoarticular, cardiovascular, neurological and genitourinary complications including epididymo-orchitis and abortion in pregnant women if left untreated [31, 32]. Main reasons for human disease are animal brucellosis in bovines and small ruminants, several risky behaviors such as consumption of unpasteurized and contaminated dairy products and not wearing protective clothing when handling potentially infectious animals and their products .
In slaughterhouse workers, villagers and farm workers of rural Pakistan, who are considered to be at special risk to get infected with brucellosis, a seroprevalence of 14% was found . These authors found a prevalence of 6.9% in a group at risk from the Potohar Plateau in 2013 . In comparable settings in India  and in Bangladesh  prevalence’s were reported to be 7.32% and 4.4%, respectively. It is also well known that a significant number of brucellosis cases can be found in patients with fever of unknown origin in endemic countries. Involving 7567 patients with suspect, yearly prevalence’s from 10.4 to 15.7% were found at a Saudi hospital between 2014 and 2018 (37). At Ayub Teaching Hospital, Abbottabad 70 patients from different districts presenting with nonspecific symptoms (fever, body aches, myalgias, arthralgia, headache, backache, malaise and insomnia) were recently screened with SAT (cutoff titer > 80) for Brucella antibodies and amazingly 49 were found positive (38). In contrast to that extraordinary high prevalence these authors documented a SAT seroprevalence of 13.6% (n = 68) in persons seeking advice at three different outdoor hospitals from Abbottabad. This prevalence shows the high burden of disease in the rural population although it could not be figured out how many admissions were brucellosis related indeed. The strongly diverging results of the two preliminary studies from Abbottabad show the need for future research to provide sustainable data for public health use. It has to be stressed that data from different studies cannot be compared without caution as various not standardized or harmonized tests are still used and those tests are not used on a routine basis at the labs either. Hence, these data show that physicians at hospitals in endemic areas should be aware of brucellosis in their day to day work.
The prevalence of brucellosis was highest in the age group 25–50 years. This finding can be explained by the fact that participants of this middle-aged group were mainly veterinarians, butchers and milking personnel who were in close contact to animals. However, seropositive cases of brucellosis were reported in participants of all age groups. As already stressed different studies cannot not be compared due to the variety of techniques used. So, the highest prevalence was found in the age group of 20–30 years (26.92%) while low prevalence was recorded in older age > 40 years (7.80%) for study participants from the Punjab, Pakistan [39 Ali 2018]. An Indian study found persons of age group 26–35 years affected most (10.8%) . A study conducted in Southern Saudi Arabia among febrile patients determined the highest seroprevalence in patients 21 and 40 years of age (35.8–45.3%), while low prevalence was recorded in young children and older people (3 and 15%), respectively . More positive cases were found in the age group of 41–80 years in Bangladesh which contrasts with our study . Hence, an interpretation of the data points to the fact, that in rural populations the presence of antibodies comes along with contact to brucellae or their LPS at work and so it is very likely to find more positives aged from 15 to 55 years. Neither the fact that anti-Brucella antibodies are detected in study participants nor that patients at hospitals are involved allow the final statement that an active infection caused those due to the shortcomings of brucellosis serology. The interpretation of these data has to be done in the light of the epidemiological context. Trends, however, are obvious and can be used to guide countermeasures.
Brucella infection was found more often in female than in male patients. Similarly, a higher prevalence of brucellosis in female patients (37%) was also recorded from Peshawar . The explanation is that animal husbandry is done mainly by women in Pakistan. Therefore, they are in direct contact to animals during their daily activities and also help during parturition without using precautionary measures. In contrast, all seropositive patients were reported to be man in a study conducted among persons of Ludhiana, India where only few women were involved in activities that exposed them to animals and other potential risk-factors . Similarly, a study conducted in high-risk group persons from Bangladesh also found a higher prevalence in man (5.6%) than in women (0.8%). The main reason for this finding was, that mainly butchers, milkers, livestock farmers and veterinary practitioners were tested. These occupations are traditionally in the hands of men there and expose them to a high risk of infection . This is also true for Egyptian setting where more man is involved in management of livestock .
This study found a higher prevalence in persons from the rural area demonstrating a higher risk of acquiring brucellosis than for people of the urban area. This finding is similar to that of a study conducted in Peshawar among hospital patients . Persons from rural areas are often involved in birthing and herding of livestock and they are more dependent on livestock production putting them at the risk of infection . A cross sectional study was conducted on rural population of the Punjab in India and seropositivity was also linked to a history assisting with abortions and calving .
Nicoletti stated that each case of brucellosis in humans is related to an animal source and its presence in animals causes a major risk of Brucella infection for humans . Thus, animal contacts, processing or handling raw animal products or meat and consumption of raw foods are the main risk factors to be considered [45, 46]. Additionally, the risk factor “access of livestock to source of drinking water” was considered to take into account epidemiological circumstances. Indeed, this study showed the highest seroprevalence in the group of persons who had direct contact with livestock (30.9%) or raised livestock at home (31.9%). As expected, processing or handling raw animal foods proved to be an important risk factor (21.3%). 28 patients recorded consumption of undercooked meat or unpasteurized milk. 13 (39.4%) participants had direct contact with materials of aborted animals. Aborted foeti are usually left for decomposition by scavengers instead of proper disposing. This procedure increases the infection risk because large numbers of organisms are excreted with the uterine fluid, placenta and fetus at the calving/lambing time .
Brucellosis in pregnant women bares the risk of abortion and may also cause repeatedly abortions after becoming chronic. Thus, participants of this study were asked about the course of previous pregnancies. Indeed, 23.8% (n = 5) seropositive women reported on abortion. A recent study in Pakistan involving 429 pregnant women mostly from rural areas reported 5.8% seroprevalence and 14.6% of these had abortion history . Due to the low number of cases in both studies, we only can recommend further studies to evaluate these findings Brucellosis can also pose a serious risk to newborns. Mortality in newborns was reported as well as transmission of brucellosis to a neonate via the congenital route or via breastmilk [48–52]. Future studies in Pakistan should also consider this neglected aspect of brucellosis in childhood. 47 reference deleted
The most common clinical sign observed among SAT seropositive patients was fever i.e. 94.1% (n = 64), followed by arthralgia with 55.8% (n = 38). Similarly, most often fever (77.71%) and arthritis (83.43%) were reported in 175 brucellosis positive hospital patients in Bikaner, India . 30.8% (n = 21) participants of this study reported persons with similar symptoms of brucellosis in their households. Transmission from person to person is rare, so these household members were most probably infected by the same animals or foods. Brucellosis in endemic countries is a family problem. Physicians should be aware of that fact and include all family members in their investigations. As brucellosis can be attracted again and again from the same source it is of imminent importance to identify and eliminate this source as well. Patients must be made aware of the epidemiology of brucellosis and local veterinary officers need to be involved finally.
RT-PCR was performed for confirmation of brucellosis in patient samples. RT-PCR is a rapid, reliable, highly sensitive and specific method for molecular diagnosis. Genus specific primers and probes targeting the conserved BCSP-31 gene were used. 11.4% samples were RT-PCR positive. The study showed that RT-PCR is a rapid method to confirm brucellosis within 2–3 hours as compared to conventional methods which take several days to weeks and also poses a high risk of infection to laboratory personnel. Cases of brucellosis might have been lost by serological tests because they were in early stage of infection and antibodies were absent still.